WO2019210194A1 - Compositions pegylées à usage oculaire et méthodes associées - Google Patents

Compositions pegylées à usage oculaire et méthodes associées Download PDF

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Publication number
WO2019210194A1
WO2019210194A1 PCT/US2019/029384 US2019029384W WO2019210194A1 WO 2019210194 A1 WO2019210194 A1 WO 2019210194A1 US 2019029384 W US2019029384 W US 2019029384W WO 2019210194 A1 WO2019210194 A1 WO 2019210194A1
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Prior art keywords
aptamer
peg
composition
attached
cases
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Christopher P. Rusconi
Carl ERICKSON
Ryan Quick
James V. MCARDLE
Matthew Levy
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Vitrisa Therapeutics Inc
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Vitrisa Therapeutics Inc
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7088—Compounds having three or more nucleosides or nucleotides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/02—Peptides of undefined number of amino acids; Derivatives thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/549—Sugars, nucleosides, nucleotides or nucleic acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/59—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes
    • A61K47/60—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0048—Eye, e.g. artificial tears
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/08—Solutions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/115—Aptamers, i.e. nucleic acids binding a target molecule specifically and with high affinity without hybridising therewith ; Nucleic acids binding to non-nucleic acids, e.g. aptamers
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/16—Aptamers
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/31—Chemical structure of the backbone
    • C12N2310/317—Chemical structure of the backbone with an inverted bond, e.g. a cap structure
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/32—Chemical structure of the sugar
    • C12N2310/321—2'-O-R Modification
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/32—Chemical structure of the sugar
    • C12N2310/322—2'-R Modification
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/35—Nature of the modification
    • C12N2310/351—Conjugate
    • C12N2310/3515—Lipophilic moiety, e.g. cholesterol

Definitions

  • Visual impairment is a national and global health concern that has a negative impact on physical and mental health.
  • Treatment of vision disorders may involve targeting certain signaling pathways or other processes in the eye.
  • intravitreal injection of one or more therapeutic agents can be used. Reducing the frequency of, or improving the safety of intravitreal injection of therapeutic agents may improve patient outcomes.
  • compositions comprising an aptamer attached or covalently bound to a polyethylene glycol (PEG) reagent comprising at least two PEG arms covalently attached to a backbone, wherein the aptamer attached or covalently bound to the PEG reagent is present in the composition at an aptamer concentration of greater than or equal to 30 mg/mL for intravitreal administration.
  • compositions for intravitreal injection comprising an aptamer attached to a PEG reagent comprising four PEG arms covalently attached to a sugar alcohol backbone.
  • the sugar alcohol backbone may comprise sorbitol, mannitol, volemitol, or any combination thereof.
  • compositions comprising an aptamer attached or covalently bound to a polyethylene glycol (PEG) reagent, wherein a given syringe format attached to a 29 or 30-gauge needle containing the composition has an injection break force of less than 13 N.
  • the aptamer may be attached or covalently bound to the PEG reagent present in the composition at an aptamer concentration of greater than or equal to 30 mg/mL.
  • the aptamer attached or covalently bound to the PEG reagent may demonstrate a statistically significant longer intravitreal half-life in a rabbit
  • the viscosity of the PEG-aptamer solution at an aptamer concentration of greater than or equal to 30 mg/mL has a viscosity such that it can be presented in a syringe format and administered by intravitreal injection at an injection break force of less than 13 N.
  • the composition may have a polydispersity index of less than 1.05.
  • the PEG reagent may comprise greater than two PEG arms attached (e.g., covalently attached) to a backbone.
  • the PEG reagent may comprise at least two PEG arms attached (e.g., covalently attached) to a backbone, wherein one or more of the at least two PEG arms has a molecular weight between 1 kDa and 10 kDa.
  • the compositions described herein may comprise a PEG reagent having a molecular weight of from at least 20 kDa to at most about 80 kDa.
  • the backbone of the PEG reagent may comprise a linear carbon backbone.
  • compositions described herein when administered via intravitreal (IVT) injection, may maintain an intravitreal concentration above 5 nM for at least 30 days in a rabbit model.
  • the aptamer comprises a nucleic acid sequence of: 5’- C 6 N H2fC m GfCfC rGf ' C m Gm GfU f ' C fU f ' C m A m Gm GfC rGf ' CfU m Gm A m GfU f ' C fU m Gm A m G fUfUrAfCfCfUmGfCmGidT-3' (SEQ ID NO: 3), wherein mG and mA are 2'O-methyl RNA; fC and fU are 2'fluoro RNA; rG and rA are 2 ⁇ H RNA; and idT is inverted deoxythymidine.
  • the aptamer comprises a nucleic acid sequence of 5'- GGUCUAGCCGGAGGAGUCAGUAAUCGGUAGACC-3' (SEQ ID NO: 5). In some embodiments, the aptamer comprises a nucleic acid sequence of 5'- GGGGGCUUAUCAUUCCAUUUAGUGUUAUGAUAACC-3' (SEQ ID NO: 4).
  • the aptamers described herein may have a Kd value of from at least 0.5 nM to at most about 10 nM.
  • Described herein are methods of treating an ocular disorder in a subject in need thereof, the methods comprising: administering a composition of any one of the preceding claims to the subject in need thereof, thereby treating the ocular disorder. Also provided herein are methods of treating an ocular disorder in a subject in need thereof, the method comprising: administering an aptamer attached to a polyethylene glycol (PEG) reagent comprising a sugar alcohol backbone to the subject in need thereof, thereby treating the ocular disorder.
  • the sugar alcohol backbone may comprise sorbitol, mannitol, volemitol, or any combination thereof.
  • PEG polyethylene glycol
  • each of the four PEG arms has a molecular weight of 10 kDa, or each of the four PEG arms has a molecular weight of less than about 10 kDa, or each of the four PEG arms has a molecular weight of less than about 5 kDa.
  • the backbone may comprise a sugar alcohol backbone.
  • the ocular disorder can be age-related macular degeneration. In some embodiments, the ocular disorder is a diabetes related eye disease, including diabetic retinopathy, peripheral diabetic retinopathy, or diabetic macular edema.
  • this disclosure provides a method of treating Stargardt disease, the method comprising: intravitreally administering an aptamer attached to a polyethylene glycol (PEG) reagent comprising four PEG arms to a subject in need thereof, thereby treating Stargardt disease, is provided.
  • PEG polyethylene glycol
  • compositions described herein may be used in treating an ocular disorder or disease.
  • the compositions described herein may be used in the formulation of a medicament for treating an ocular disease.
  • the methods described herein comprise administering the aptamer covalently bound to a polyethylene glycol (PEG) reagent by intravitreal injection via a syringe to the subject in need.
  • PEG polyethylene glycol
  • the syringe is attached to a 27-gauge needle.
  • the syringe is attached to a 28-gauge needle. In some cases, the syringe is attached to a 29-gauge needle. In some cases, the syringe is attached to a 30-gauge needle. In some cases, the syringe is attached to a 3 l-gauge needle. In some cases, the syringe is attached to a needle with a 27-gauge or greater. In some cases, the syringe is attached to a needle with a 28-gauge or greater. In some cases, the syringe is attached to a needle with a 29-gauge or greater. In some cases, the syringe is attached to a needle with a 30-gauge or greater. In some cases, the syringe is attached to a needle with a 31 -gauge or greater.
  • the present disclosure overcomes some challenges involved in injecting therapeutics into delicate tissues (e.g., eye tissue) that are susceptible to damage when encountering a relatively high level of injection break force.
  • the present disclosure provides PEGylated aptamers that can be delivered intravitreally at higher aptamer concentrations in a single administration to provide a greater administered dose.
  • formulations containing the PEGylated aptamers provided herein have reduced viscosity and thus the PEGylated aptamers can be delivered with reduced levels of injection break force.
  • the PEGylated aptamers may have superior therapeutic efficacy compared to aptamers that need to be delivered at lower doses in order to minimize viscosity.
  • the PEGylated aptamers provided herein may also have longer intravitreal residence times, which may also positively impact their therapeutic efficacy.
  • FIG. 1 provides non-limiting examples of chemical structures PEG reagents in accordance with various aspects of the disclosure.
  • FIG 2. provides a non-limiting example of the relationship between viscosity and concentration for several PEG-aptamers of the disclosure. Injection break force limits for IVT injection via a 29G or 30G needle in combination with a suitable syringe presentation as calculated per Equation 1 of the disclosure are shown.
  • FIG. 3 provides illustrative formulations and pharmacokinetics of PEGylated aptamers in accordance with various aspects of the disclosure.
  • the PEGylated aptamers provided herein may comprise an aptamer attached to one or more PEG reagents that have branching configurations.
  • the PEGylated aptamers contain PEG reagents with multiple PEG arms, such as at least two PEG arms, at least four PEG arms, at least six PEG arms, at least eight arms or higher numbers of PEG arms.
  • one or more PEG arms within the PEG reagent has a molecular weight of about lOldDa or less.
  • the PEGylated aptamers described herein can comprise a PEG reagent with PEG arms attached to a certain type of backbone, such as a sugar alcohol backbone.
  • the PEGylated aptamers may comprise a PEG reagent with four PEG arms covalently attached to a sugar alcohol backbone.
  • This disclosure also provides formulations containing such PEGylated aptamers, particularly formulations with relatively high aptamer concentrations (e.g ., greater than 30 mg/mL, greater than about 35 mg/mL, greater than about 40 mg/mL, greater than about 45 mg/mL, greater than about 50 mg/mL, or more based on the molecular weight of the aptamer moiety only) that yet maintain a viscosity that enables the aptamers to be injectable, particularly in the setting of the eye or other delicate tissue.
  • relatively high aptamer concentrations e.g ., greater than 30 mg/mL, greater than about 35 mg/mL, greater than about 40 mg/mL, greater than about 45 mg/mL, greater than about 50 mg/mL, or more based on the molecular weight of the aptamer moiety only
  • a formulation provided herein may require a relatively low injection force (e.g., less than about 13 N, or less than about 10 N) when injected through a syringe with a needle suitable for use for intravitreal administration (e.g, a 29 or 30-gauge needle).
  • a needle suitable for use for intravitreal administration e.g, a 29 or 30-gauge needle.
  • the PEGylated aptamers provided herein may have other features, such as relatively long intravitreal half-lives (e.g, greater than about 5 days in rabbits, or greater than about 10 days in humans).
  • This disclosure also provides for formulations containing such PEGylated aptamers, particularly formulations with aptamer concentrations greater than 30 mg/mL suitable for intravitreal administration to the eye.
  • the eye is a fragile tissue, which limits both the size of the needle that can be used as well as the injection, or break force, applied to the plunger for intravitreal administration of drugs to the eye.
  • Break force is the viscous resistive force required to move the plunger of the syringe, measured in Newtons (N), or in one aspect, the force required to initiate movement of the plunger.
  • the maximum tolerable break force is accepted as less than or equal to approximately 12 N, or less than 13 N (Martinez-Sancho, C., Herrero-Vanrell, R., and Negro, S. (2004). J. Controlled Release 99: 41-52; Martinez-Sancho, C., Herrero-Vanrell, R., and Negro, S. (2006). Int J. Pharmaceutics 326: 100-106).
  • R b Barrel radius (mm)
  • the total injection break force on the plunger is equal to the sum of the viscous resistive force and the friction related to moving the piston. Since in most syringes, there is technology to mitigate this friction, one can assume that this force is negligible when compared to the viscous resistive force.
  • the injection force (break force) may be calculated to be equal to the viscous resistive force, which can be estimated from equation 1.
  • equation 1 can be modeled once a syringe/needle combination is selected. With that combination, the injection break force can be estimated for a wide variety of materials for which viscosity data exists.
  • the branching configuration of the PEG reagent may influence the viscosity properties and/or maximum formulation strength suitable for intravitreal administration, as defined by an injection break force of less than 13 N.
  • FIG. 2 provides viscosity versus concentration data for three representative PEG reagents of the same molecular weight of about 40 kDa conjugated to the same aptamer, but with different branching configurations and with markedly different properties, with injection break force limits for syringes of a given configuration using a 29G or 30G needle.
  • attaching a PEG reagent with a higher number of PEG arms in the molecule may increase the aptamer concentration of PEGylated aptamer that may be used in a formulation suitable for ocular delivery (and thus, improve the therapeutic efficacy of the formulation) as compared to attaching the same aptamer to a PEG reagent with a lower number of PEG arms in the molecule.
  • attaching a PEG reagent with a higher number of PEG arms in the molecule may increase the aptamer concentration of PEGylated aptamer that may be used in a formulation suitable for ocular delivery with a higher gauge needle (and thus, improve the tolerability and safety of the injection), as compared to attaching the same aptamer to a PEG reagent with a lower number of PEG arms in the molecule.
  • the PEGylated aptamers provided herein may have superior properties for intravitreal administration when compared with PEGylated aptamers with a similar molecular weight but having a different branching configuration.
  • PEGylated aptamers with greater numbers of PEG arms may have superior viscosity profiles than PEGylated aptamers with fewer numbers of PEG arms (e.g., 2 PEG arms or less).
  • formulations containing PEGylated aptamers with four PEG arms may maintain intravitreal injectability at higher aptamer concentrations compared with PEGylated aptamers with two PEG arms and the same molecular weight of PEG.
  • Aptamer compositions with higher numbers of PEG arms may have greater intravitreal half- life compared with aptamers with fewer numbers of PEG arms and may require less injection force during delivery.
  • PEGylated aptamers comprising a PEG reagent with about four PEG arms can comprise an aptamer concentration greater than about 50 mg/mL
  • PEGylated aptamers comprising a PEG reagent of the same molecular weight with about two PEG arms can comprise an aptamer concentration of about at most 30 mg/mL.
  • PEGylated aptamers with 1 or 2 PEG arms may also be useful. As such, in some
  • this disclosure provides PEGylated aptamers with 1 PEG arm, 2 PEG arms, or 2 or more PEG arms.
  • this disclosure provides PEGylated aptamers for intravitreal administration comprising a PEG reagent with at least two arms or at least three arms, or more, and can be formulated to have an aptamer concentration greater than about 30 mg/mL, greater than about 35 mg/mL, greater than about 40 mg/mL, greater than about 45 mg/mL, or greater than about 50 mg/mL, or more.
  • an aptamer concentration greater than about 30 mg/mL, greater than about 35 mg/mL, greater than about 40 mg/mL, greater than about 45 mg/mL, or greater than about 50 mg/mL, or more.
  • PEGylated aptamers provided herein a 28-, 29-, 30- or 31 -gauge needle (or other type of needle suitable for ocular or intravitreal injection) containing the PEGylated aptamers provided herein may be used with an injection break force that is relatively low, such as less than about 13 N as described in more detail below.
  • formulations and compositions provided herein can have several benefits as therapeutic agents for diseases of the retina and/or diseases treatable by intravitreal administration of a therapeutic.
  • formulations and compositions of PEGylated aptamers comprising PEG reagents with a greater branching configuration e.g., 2 or more PEG arms, 3 or more PEG arms
  • a greater branching configuration e.g., 2 or more PEG arms, 3 or more PEG arms
  • each dose can deliver a higher aptamer concentration of therapeutic aptamers compared with each dose for PEGylated aptamers comprising PEG reagents of the same molecular weight with less branching configuration.
  • formulations and compositions can be delivered to the eye ( e.g ., via intravitreal administration) and can be especially suited for delivery to relatively delicate or fragile areas of the body.
  • formulations with relatively low viscosity can be delivered using higher gauge needles (e.g., 28-, 29-, 30-, 3 l-gauge needles). This improves patient comfort with the therapy and is likely to enhance patient adherence to the therapy.
  • Use of higher gauge needles also improves the safety of therapeutic delivery and reduces the risk of injury and/or infection, especially to relatively delicate or fragile areas of the body such as the eye. Less force may be applied to the body surfaces in order to deliver the therapeutic
  • composition thereby reducing the likelihood and/or risk of injuries.
  • Improved safety can be assessed in a number of ways.
  • enhanced safety results in a lower incidence of elevated intraocular pressure (e.g., about 30 minutes or about 60 minutes following intravitreal injection), a lower incidence of endophthalmitis, retinal detachment, iatrogenic traumatic cataract, or a combination thereof, following intravitreal injection.
  • enhanced safety reduces the number of reported ocular adverse reactions such as conjunctival hemorrhage, conjunctivitis, eye pain, vitreous floaters, intraocular inflammation, cataract, foreign body sensation in eyes, eye irritation, increase in lacrimation, blepharitis, dry eye, visual disturbance or vision blurring, eye pruritis, ocular hyperemia, retinal disorder, maculopathy, retinal degeneration, ocular discomfort, conjunctival hyperemia, posterior capsule opacification, injection site hemorrhage, or a combination thereof.
  • ocular adverse reactions such as conjunctival hemorrhage, conjunctivitis, eye pain, vitreous floaters, intraocular inflammation, cataract, foreign body sensation in eyes, eye irritation, increase in lacrimation, blepharitis, dry eye, visual disturbance or vision blurring, eye pruritis, ocular hyperemia, retinal disorder, maculopathy, retinal degeneration, ocular discomfort, conjunctival hyperemia, posterior
  • the ocular disease is Stargardt disease.
  • the ocular disease or disorder is a macular degeneration such as, for example, age-related macular degeneration (AMD) or juvenile macular degeneration. AMD includes, but is not limited to dry AMD, or neovascular (Wet) AMD.
  • the ocular disease is a diabetes related eye disease, including but not limited to diabetic retinopathy (DR), peripheral diabetic retinopathy (PDR) or diabetic macular edema (DME).
  • DR diabetic retinopathy
  • PDR peripheral diabetic retinopathy
  • DME diabetic macular edema
  • the ocular disease is a macular edema; macular edema includes, but is not limited to, retinal vein occlusion (RVO).
  • ocular diseases and disorders include, but are not limited to, Polyploidal Choroidal Vasculopathy (PCV), Myopic Choroidal Neovascularization (mCNV), Proliferative Vitreoretinopathy, Glaucoma, Uveitis, Keratitis, ocular hypertension, a retinoblastoma, Blepharitis, Nystagmus, Retinitis, a subconjunctive hemorrhage, or a combination thereof.
  • PCV Polyploidal Choroidal Vasculopathy
  • mCNV Myopic Choroidal Neovascularization
  • aptamer refers to oligonucleotide molecules that bind to a target (e.g a protein) with high affinity and specificity through non-Watson-Crick base pairing interactions.
  • a target e.g a protein
  • the aptamers described herein are non-naturally occurring oligonucleotides (i.e., synthetically produced) that are isolated and used for the treatment of a disorder or a disease.
  • Aptamers can bind to essentially any target molecule including, without limitation, proteins, oligonucleotides, carbohydrates, lipids, small molecules, and even bacterial cells.
  • the aptamers described herein are oligonucleotides that bind to a target or inhibit a function associated with a target. Whereas many naturally occurring
  • oligonucleotides such as mRNA
  • aptamers can be distinguished from these naturally occurring oligonucleotides in that binding of the aptamer to a target molecule is dependent upon secondary and tertiary structures of the aptamer rather than a conserved linear base sequence.
  • the aptamer generally does not encode information in its linear base sequence.
  • the term“or” is used nonexclusively to encompass“or” and“and.”
  • “A or B” includes“A but not B,”“B but not A,” and“A and B” unless otherwise indicated.
  • sequence identity refers to an exact nucleotide-to-nucleotide or amino acid-to-amino acid correspondence of two polynucleotides or polypeptide sequences, respectively.
  • techniques for determining sequence identity include determining the nucleotide sequence of a polynucleotide and/or determining the amino acid sequence encoded thereby, and comparing these sequences to a second nucleotide or amino acid sequence.
  • Two or more sequences can be compared by determining their“percent identity.”
  • the percent identity of two sequences, whether nucleic acid or amino acid sequences is the number of exact matches between two aligned sequences divided by the length of the shorter sequence and multiplied by 100. Percent identity may also be determined, for example, by comparing sequence information using the advanced BLAST computer program, including version 2.2.9, available from the National Institutes of Health.
  • the BLAST program is based on the alignment method of Karlin and Altschul, Proc. Natl. Acad. Sci. USA, 87:2264-2268 (1990) and as discussed in Altschul, et al., J. Mol.
  • the program may be used to determine percent identity over the entire length of the oligonucleotides or proteins being compared. Default parameters are provided to optimize searches with short query sequences in, for example, with the blastp program.
  • the program also allows use of an SEG filter to mask-off segments of the query sequences as determined by the SEG program of Wootton and Federhen, Computers and Chemistry, 17: 149-163 (1993).
  • Ranges of desired degrees of sequence identity are approximately 80% to 100% and integer values therebetween.
  • this disclosure encompasses sequences with at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, or at least 98% sequence identity with any sequence provided herein.
  • “modification identity” refers to two polynucleotides with identical patterns of modifications on a nucleotide-to-nucleotide level.
  • Techniques for determining modification identity may include determining the modifications of a polynucleotide and comparing these modifications to modifications of a second polynucleotide.
  • the percent modification identity of two sequences is the number of exact modification matches between two aligned sequences divided by the length of the longer sequence and multiplied by 100. Ranges of desired degrees of modification identity are generally approximately 50% to 100%.
  • this disclosure encompasses sequences with at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 98% modification identity with any sequence provided herein.
  • compositions that comprise one or more aptamers attached to one or more polyethylene glycol (PEG) reagents, referred to herein as PEGylated aptamers.
  • PEG polyethylene glycol
  • the PEG reagents described herein are linear, branched, networked, cross-linked, or a combination thereof.
  • the PEG reagents are branched.
  • the term“PEG” generally refers to a polymer or oligomer containing repeating units of ethylene oxide (O-CEL-CED n ⁇
  • the PEG reagents described herein contain a simple chemical backbone, such as a carbon backbone, that is attached to 2 or more PEG arms (or branches or chains).
  • the term“arm” as used herein refers to a linear PEG that is attached to another molecule, such as a backbone or a PEG branch. As used herein, the term“arm” does not refer to a linear PEG that is directly attached to multiple linear PEGs.
  • a “branch” is structurally similar to a PEG arm in that it may comprise repeating units of ethylene oxide and is also linear in structure. A branch may be identical in structure to an arm and, like an arm, may be attached to a backbone of the molecule.
  • a“PEG branch” also describes a linear PEG molecule that is attached to one or more linear PEGs, such as one or more PEG arms or one or more PEG branches.
  • the PEG reagents provided herein may contain multiple PEG arms and/or branches.
  • a PEG reagent provided herein may comprise a backbone attached to a single PEG branch, which is attached to two or more PEG arms ( e.g.,2 arms, 3 arms, 4 arms, 5 arms, 6 arms, 7 arms, 8 arms or more).
  • a PEG reagent provided herein may comprise a backbone attached to two or more PEG branches; and at least one of the branches may be attached to 2 or more PEG arms.
  • the number of arms directly attached or indirectly attached (e.g ., attached via a branch or other structure) to the backbone of the PEG reagent can vary.
  • the PEG reagent can comprise 2 or more arms (or chains) attached to the linear backbone.
  • the PEG reagent can comprise multiple arms (e.g., multiple PEG arms), such as at least 2 arms, at least 3 arms, at least 4 arms, at least 5 arms, at least 6 arms, at least 7 arms, at least 8 arms, at least 9 arms, or at least 10 arms.
  • the PEG reagent can comprise less than 3 arms, less than 4 arms, less than 5 arms, less than 6 arms, less than 7 arms, less than 8 arms, less than 9 arms, or less than 10 arms.
  • the 3 or more arms (or chains) can comprise PEG.
  • the PEG reagent may have at least three PEG arms. In some instances, the PEG reagent may have at least four PEG arms. In some instances, the PEG reagent may contain at least five PEG arms. In some instances, the PEG reagent may contain at least six PEG arms. In some instances, the PEG reagent may contain at least six PEG arms. In some instances, the PEG reagent may contain at least seven PEG arms.
  • the PEG reagent may contain at least eight PEG arms. In some instances, the PEG reagent may contain at least nine PEG arms. In some instances, the PEG reagent may contain at least ten PEG arms. In some instances, the PEG reagent may contain at least eleven PEG arms. In some instances, the PEG reagent may contain at least twelve PEG arms. In some instances, the PEG reagent may contain at least thirteen PEG arms. In some instances, the PEG reagent may contain at least fourteen PEG arms. In some instances, the PEG reagent may contain at least fifteen PEG arms. In some instances, the PEG reagent may contain at least sixteen PEG arms. In some instances, the PEG reagent may contain at least seventeen PEG arms.
  • the PEG reagent may contain at least eighteen PEG arms. In some instances, the PEG reagent may contain at least nineteen PEG arms. In some instances, the PEG reagent may contain at least twenty PEG arms. In some cases, a PEG reagent contains 2-20 PEG arms.
  • the number of branches directly attached or indirectly attached to the backbone of the PEG reagent can vary.
  • the PEG reagent can comprise 2 or more branches or 3 or more branches attached to the linear backbone.
  • the PEG reagent can comprise 2 branches, 3 branches, 4 branches, 5 branches, 6 branches, 7 branches, 8 branches, 9 branches, or 10 branches.
  • the 2 or more branches can comprise PEG.
  • the PEG reagent may have at least three PEG branches.
  • the PEG reagent may have at least four PEG branches.
  • the PEG reagent may contain at least six PEG branches.
  • a PEG reagent contains 2-20 PEG branches.
  • the length of a PEG branch defined as the molecular weight or number of ethylene oxide units can vary. In some instances, shorter PEG branches provide for pegylation reagents that when conjugated to an aptamer, result in formulations with lower viscosity at any given PEG-aptamer formulation strength. Not wishing to be bound by theory, when the arm length of a pegylation reagent is shorter, the likelihood of inter-strand interactions between PEG arms of PEG-conjugated molecules is less likely, and therefore, the viscosity of the solution may be lower. This property is independent of the number of PEG arms of a pegylation reagent.
  • pegylation reagents with a greater number of arms may, when conjugated to an aptamer, result in PEG- aptamer formulations with a lower viscosity as compared to PEG-aptamers comprising PEGs of fewer arms.
  • the PEG reagent comprises a linear backbone.
  • the PEG reagent can comprise a linear carbon backbone.
  • the linear carbon backbone can comprise at least 2 carbon atoms, at least 3 carbon atoms, at least 4 carbon atoms, at least 5 carbon atoms, at least 6 carbon atoms, at least 7 carbon atoms, at least 8 carbon atoms, at least 9 carbon atoms, or at least 10 carbon atoms.
  • the linear carbon backbone may also contain constituents other than carbon.
  • the linear carbon backbone can comprise a sugar alcohol.
  • the sugar alcohol comprises sorbitol, mannitol, glycerol, ribitol, xylitol, arabitol, erythritol, threitol, inositol, volemitol, etc.
  • a PEG reagent with a sugar alcohol backbone may have improved properties compared to a PEG reagent without a sugar alcohol.
  • a PEGylated aptamer using a PEG reagent with a sugar alcohol backbone may have a longer half-life than a PEGylated aptamer containing a PEG reagent without a sugar alcohol backbone.
  • the half-life of the PEGylated aptamer is at least 1.5 fold, at least 2-fold, at least 2.5-fold, at least 3 fold, at least 3.5 fold, at least 4 fold, at least 5 fold, at least 10 fold longer than a PEGylated aptamer containing a PEG reagent that lacks sugar alcohol.
  • FIG. 1 provides some exemplary PEG reagent structures that may be attached to an aptamer, as provided herein.
  • PEG Reagent 1 (FIG. 1 A) provides an exemplary structure of a PEG reagent with two arms, arranged such that both the first and second arms are directly attached to the backbone ( e.g ., carbon backbone).
  • the PEG reagent structures attached to the aptamers herein may include PEG reagents with similar structures as PEG Reagent 1 but arranged in a different configuration as shown, for example, in FIG. 1B and FIG. 1C.
  • a PEG reagent may comprise two or more PEG arms directly attached to a backbone (e.g., carbon backbone).
  • a PEG reagent may comprise at least 4, at least 6, at least 8, or more PEG arms directly attached to the backbone (e.g, carbon backbone).
  • the PEG reagent has four PEG arms, two of which are attached to one PEG branch of the molecule and two of which are attached to a second PEG branch of the molecule.
  • each branch in the PEG reagent is attached to two PEG arms.
  • at least one of the branches in the PEG reagent is attached to at least two PEG arms, such as three or more PEG arms, four or more PEG arms, or more.
  • at least one of the branches in the PEG reagent is not attached to a PEG arm.
  • the PEG reagent has four PEG arms and a carbon alcohol linear backbone of about five (5) carbons.
  • each PEG arm is attached to an oxygen atom on the carbon alcohol backbone.
  • these PEG-arm-attached carbon atoms are directly linked to each other. However, in some embodiments, there may be intervening components between the carbon atoms (or other atom) attached to the PEG arms.
  • the PEGylated aptamers provided herein may contain an aptamer attached to a PEG reagent at its 5' end or its 3' end.
  • the PEG reagent may be attached to a nucleotide of an aptamer that is not located at its 5' or 3' end.
  • the aptamer can be attached to the PEG reagent via an ionic bond or a covalent bond.
  • the therapeutic composition comprises an aptamer covalently bound to a PEG reagent.
  • the aptamer can be attached directly to a PEG reagent. In some cases, the aptamer can be attached indirectly to a PEG reagent through a linker.
  • the linker can be a functional group of the PEG reagent described above or of the aptamer.
  • the aptamer can be functionalized with a linker that contains a unique reactive moiety such as a primary amine or thiol group.
  • a PEGylation reagent can contain a leaving group, such as an NHS or maleimide moiety attached to the PEG reagent via a linker.
  • the aptamer can be attached to a PEG reagent directly or with the use of a spacer or linker.
  • a spacer or linker In a non-limiting example, 6-(trifluoroacetamido)hexanol (2- cyanoethyl-N,N-diisopropyl)phosphoramidite can be used to add a hexylamino linker to the 5' end of the synthesized aptamer.
  • This linker as with the other amino linkers provided herein, once the group protecting the amine has been removed, can be reacted with PEG-NHS esters to produce covalently linked PEG-aptamers.
  • Other non-limiting examples of linker phosphoramidites may include: TFA-amino C4 CED phosphoramidite having the structure:
  • 5'-amino modifier 5 having the structure:
  • the 5'-thiol modified linker may be used, for example, with PEG-maleimides, PEG- vinylsulfone, PEG-iodoacetamide and PEG-orthopyridyl-disulfide.
  • the aptamer may be bonded to the 5'-thiol through a maleimide or vinyl sulfone functionality.
  • FIG. 1 provides exemplary PEG reagent structures of the same total molecular weight of about 40kDa.
  • PEG reagent 1 comprises a 40 kDa PEG reagent with two PEG arms of about 20 kDa each (FIG. 1A).
  • PEG reagent 2 comprises four PEG arms, each about 7.5 kDa (FIG. IB).
  • PEG reagent 3 also comprises four (4) PEG arms, each arm about 10 kDa (FIG. 1C)
  • the molecular weight of one or more PEG arms in a PEGylated aptamer provided herein is from about 1 kDa to about 30 kDa. In some cases, the molecular weight of each arm comprising PEG is at least about 1 kDa. In some cases, the molecular weight of each arm comprising PEG is less than about 30 kDa. In some cases, the molecular weight of each arm comprising PEG is about 5 kDa, about 10 kDa, about 20 kDa, or about 25kDa.
  • the molecular weight of each arm comprising PEG is from about 5 kDa to about 10 kDa, from about 10 kDa to about 15 kDa, from about 15 kDa to about 20 kDa, or from about 20 kDa to about 30 kDa.
  • the total molecular weight of a PEG reagent provided herein can, in some cases, be from about 20 kDa to about 80 kDa. In some cases, the total molecular weight of the PEG reagent is at least about 20 kDa. In some cases, the total molecular weight of the PEG reagent is at most about 80 kDa. In some cases, the total molecular weight of the PEG reagent is from about 20 kDa to about 30 kDa.
  • the total molecular weight of the PEG reagent is from about 30 kDa to about 40 kDa, from about 40 kDa to about 50 kDa, from about 50 kDa to about 60 kDa, from about 60 kDa to about 70 kDa, or from about 70 kDa to about 80 kDa. In some cases, the total molecular weight of the PEG reagent is about 35 kDa, about 40 kDa, about 45 kDa, about 50 kDa, about 55 kDa, about 60 kDa, about 65 kDa, about 70 kDa, or about 75 kDa.
  • the PEG reagent can comprise one or more functional groups.
  • the functional group can comprise, for example, a carboxylic acid, an ester, a carbonate, an aldehyde, an amine, an oxyamine, a hydrazide, an azide, an unsaturated bond, a thiol, a dithiopyridine, a sulfone, a maleimide, a vinylsulfone, an a-iodoacetyl, an acrylate, an isocyanate, an isothiocyanate, an epoxide, reagents thereof and/or combinations thereof.
  • the PEGylated aptamers described herein may comprise one or more aptamers that comprise a DNA oligonucleotide, an RNA oligonucleotide, a modified DNA, a modified RNA oligonucleotide, or a combination thereof.
  • the aptamer can be an oligonucleotide molecule that binds to a target with high affinity and specificity through non-Watson-Crick base pairing interactions.
  • the aptamer can comprise naturally-occurring nucleotide bases, non-naturally occurring nucleotide bases, or a combination thereof.
  • the aptamer can bind to different types of targets including, without limitation, proteins, peptides, oligonucleotides, carbohydrates, lipids, small molecules, and microbial cells such as bacteria.
  • an aptamer is isolated or purified.“Isolated” (used interchangeably with “substantially pure” or“purified”) as used herein means that an aptamer that is synthesized chemically; or has been separated from other aptamers. It will be understood that some compositions can contain a mixture of aptamers.
  • the length of the aptamer can be variable. In some cases, the length of the aptamer is less than 100 nucleotides. In some cases, the length of the aptamer is greater than 10 nucleotides. In some cases, the length of the aptamer is between 10 and 90 nucleotides.
  • the aptamer can be, without limitation, about 10, about 15, about 20, about 25, about 30, about 35, about 40, about 45, about 50, about 55, about 60, about 65, about 70, about 75, about 80, about 85, or about 90 nucleotides in length.
  • the aptamer can comprise from about 20 nucleotides to about 40 nucleotides. In some cases, the aptamer comprises at least about 20 nucleotides. In some cases, the aptamer comprises at most about 40 nucleotides. In some instances, the aptamer comprises from 28 to 40 nucleotides. In some instances, the aptamer comprises from 37 to 46 nucleotides.
  • aptamers can be modified, without limitation, to provide other chemical groups that incorporate additional charge, polarizability, hydrophobicity, hydrogen bonding, electrostatic interaction, and functionality to the nucleic acid aptamer bases or to the nucleic acid aptamer as a whole.
  • Modifications to generate oligonucleotide populations that are resistant to nucleases can also include, for example, one or more substitute intemucleotide linkages, altered sugars, altered bases, or combinations thereof.
  • Such modifications include, but are not limited to, 2'-position sugar modifications, 5-position pyrimidine modifications, 8- position purine modifications, modifications at exocyclic amines, substitution of 4- thiouridine, substitution of 5-bromo or 5-iodo-uracil; backbone modifications,
  • Modifications can also include 3' and 5' modifications such as capping, e.g., addition of a 3'- 3'-dT cap to increase exonuclease resistance.
  • modifications may include chemical substitutions at the sugar and/or phosphate and/or base positions, for example, at the 2' position of ribose, the 5' position of pyrimidines, and the 8' position of purines, various 2'-modified pyrimidines and modifications with 2'-amino (2 -NH2), 2'-fluoro (2'-F), and/or 2'-0-methyl (2'-OMe) substituents.
  • aptamers described herein comprise a 2'-OMe and/or a 2'F modification to increase in vivo stability.
  • the aptamers described herein contain modified nucleotides to improve the affinity and specificity of the aptamers for a specific epitope, exosite or active site.
  • modified nucleotides include those modified with guanidine, indole, amine, phenol, hydroxymethyl, or boronic acid.
  • pyrimidine nucleotide triphosphate analogs or CE-phosphoramidites may be modified at the 5 position to generate, for example, 5-benzylaminocarbonyl-2’-deoxyuridine (BndET); 5-[N-(phenyl-3- propyl)carboxamide]-2'-deoxyuridine (PPdET); 5-(N-thiophenylmethylcarboxyamide)-2'- deoxyuridine (ThdET); 5-(N-4-fluorobenzylcarboxyamide)-2'-deoxyuridine (FBndET); 5-(N- (l-naphthylmethyl)carboxamide)-2 " -deoxyuridine (NapdET); 5 -(N-2- naphthylmethylcarboxyamide)-2'-deoxyuridine (2NapdU); 5-
  • Modifications of the aptamers contemplated in this disclosure include, without limitation, those which provide other chemical groups that incorporate additional charge, polarizability, hydrophobicity, hydrogen bonding, electrostatic interaction, and functionality to the nucleic acid aptamer bases or to the nucleic acid aptamer as a whole.
  • Modifications to generate oligonucleotide populations that are resistant to nucleases can also include one or more substitute internucleotide linkages, altered sugars, altered bases, or combinations thereof.
  • modifications include, but are not limited to, 2'-position sugar modifications, 5-position pyrimidine modifications, 8-position purine modifications, modifications at exocyclic amines, substitution of 4-thiouridine, substitution of 5-bromo or 5-iodo-uracil; backbone modifications, phosphorothioate, phosphorodithioate, or alkyl phosphate modifications, methylations, and unusual base-pairing combinations such as the isobases isocytidine and isoguanosine.
  • Modifications can also include 3' and 5' modifications such as capping, e.g., addition of a 3'-3'-dT cap to increase exonuclease resistance.
  • Aptamers of the disclosure may generally comprise nucleotides having ribose in the b-D-ribofuranose configuration. In some cases, 100% of the nucleotides present in the aptamer have ribose in the b-D-ribofuranose configuration. In some cases, at least 50% of the nucleotides present in the aptamer have ribose in the b-D-ribofuranose configuration. In some cases, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or 100% of the nucleotides present in the aptamer have ribose in the b-D-ribofuranose configuration.
  • the therapeutic composition comprises a PEGylated aptamer in which the aptamer binds to and inhibits complement Factor D (fD) protein.
  • the aptamer binds to vascular endothelial growth factor (VEGF).
  • VEGF vascular endothelial growth factor
  • the aptamer binds to angiopoietin-2 (Ang2).
  • the aptamer binds to complement C5a.
  • the aptamer binds to interleukin-8 (IL-8).
  • the aptamer binds to interleukin- 17 (IL-17).
  • a PEGylated aptamer to be used in the methods described herein comprises any one of the aptamer sequences listed in Table 1 :
  • the branching configuration of the PEG reagent may influence the pharmacokinetics and/or pharmacodynamics of the therapeutic composition and/or the viscosity properties of the therapeutic composition in suitable formulation buffers.
  • FIG. 3 provides three representative PEG reagents of the same molecular weight of about 40 kDa but with different branching configurations and with markedly different properties.
  • attaching a PEG reagent with a higher number of PEG arms in the molecule may increase the aptamer concentration of PEGylated aptamer that may be used in a formulation suitable for ocular delivery (and thus, improve the therapeutic efficacy of the formulation) as compared to attaching the same aptamer to a PEG reagent with a lower number of PEG arms in the molecule.
  • attaching a PEG reagent with a higher number of PEG arms in the molecule may increase the intravitreal half-life of the PEGylated aptamer as compared to attaching the same aptamer to a PEG reagent with a lower number of PEG arms in the molecule.
  • this disclosure provides PEGylated aptamers for intravitreal administration that contain an aptamer attached to a PEG reagent with multiple arms and that also have a maximum formulation viscosity above a certain value.
  • this disclosure provides PEGylated aptamers attached to a PEG reagent that contains at least two arms and the PEGylated aptamer is formulated at an aptamer concentration greater than about 30 mg/mL, greater than about 35 mg/mL, greater than about 40 mg/mL, greater than about 45 mg/mL, greater than about 50 mg/mL, or higher.
  • such PEGylated aptamers are injectable, e.g., injectable via a needle suitable for intra-ocular or intravitreal injection, when formulated at an aptamer concentration greater than about 30 mg/mL, greater than about 35 mg/mL, greater than about 40 mg/mL, greater than about 45 mg/mL, greater than about 50 mg/mL, or higher.
  • this disclosure provides PEGylated aptamers attached to a PEG reagent that contains at least four arms and the PEGylated aptamers are present in a formulation at an aptamer concentration greater than about 30 mg/mL, greater than about 35 mg/mL, greater than about 40 mg/mL, greater than about 45 mg/mL, greater than about 50 mg/mL, or higher.
  • such PEGylated aptamers are injectable, e.g, injectable via a needle suitable for intra-ocular or intravitreal injection, when formulated at an aptamer concentration greater than about 30 mg/mL, greater than about 35 mg/mL, greater than about 40 mg/mL, greater than about 45 mg/mL, greater than about 50 mg/mL, or higher.
  • the PEGylated aptamer is formulated at a total (weight inclusive of the molecular weight of both the aptamer and PEG moeity) concentration greater than about 150 mg/mL, greater than about 175 mg/mL, greater than about 200 mg/mL, greater than about 225 mg/mL, greater than about 250 mg/mL, or higher.
  • such PEGylated aptamers are injectable, e.g, injectable via a syringe/needle presentation suitable for intra- ocular or intravitreal injection, when formulated at a total (aptamer and PEG reagent) concentration greater than about 150 mg/mL, greater than about 175 mg/mL, greater than about 200 mg/mL, greater than about 225 mg/mL, greater than about 250 mg/mL, or higher.
  • this disclosure provides PEGylated aptamers attached to a PEG reagent that contains at least four arms and the PEGylated aptamers are present in a formulation at a total (aptamer and PEG reagent) concentration greater than about 150 mg/mL, greater than about 175 mg/mL, greater than about 200 mg/mL, greater than about 225 mg/mL, greater than about 250 mg/mL, or higher.
  • such PEGylated aptamers are injectable, e.g, injectable via a syringe/needle presentation suitable for intra-ocular or intravitreal injection, when formulated at a total (aptamer and PEG reagent) concentration greater than about 150 mg/mL, greater than about 175 mg/mL, greater than about 200 mg/mL, greater than about 225 mg/mL, greater than about 250 mg/mL, or higher.
  • the increased branching or increasing the number of PEG arms in a PEG reagent not only affects the maximum aptamer concentration while maintaining injectability.
  • a PEGylated aptamer comprising PEG reagent 1 having two (2) PEG arms may have an intravitreal half-life of about 110 hours in rabbits.
  • the intravitreal half-life may increase to about 130 hours in rabbits.
  • this disclosure provides PEGylated aptamers containing PEG reagents with at least two arms or at least three arms, or more, and that have an intravitreal half-life of at least about 4.0 days, at least about 4.5 days, at least about 5.0 days, at least about 5.5 days, at least or about 6.5, at least or about 7.5 days or greater, following intravitreal injection of rabbits.
  • the PEGylated aptamers described herein may have an intravitreal half-life of at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, or greater, following intravitreal injection of, for example, a human subject.
  • PEGylated aptamers containing PEG reagents with three or more branches (or arms) may support improved therapeutic formulations for intravitreal administration because such therapeutic formulations may have a higher aptamer concentration of PEGylated aptamer, while still maintaining injectability suitable for the eye.
  • Formulations with relatively high aptamer concentrations and relatively low viscosity may allow for delivery to the eye of higher aptamer concentrations per dose.
  • compositions that can be delivered at relatively lower intravitreal dosing intervals (e.g once every 6 weeks, once every 8 weeks, once every 12 weeks, once every 16 weeks, once every 20 week, once every 24 weeks, etc.) in order to achieve a desired therapeutic effect.
  • relatively lower intravitreal dosing intervals e.g once every 6 weeks, once every 8 weeks, once every 12 weeks, once every 16 weeks, once every 20 week, once every 24 weeks, etc.
  • smaller volumes can be delivered to relatively fragile parts of the body, such as the eye, which improves the safety of the therapy.
  • Low viscosity may allow for better injectability with clinically preferred high gauge needles.
  • These properties provide several benefits including increased therapeutic effect due to the delivery of higher concentrations/dose; reduced risk of injury due to fewer injections required; and enhanced commercial and safety properties due to the use of high gauge needles.
  • the PEGylated aptamers can have enhanced pharmacokinetics and efficacy with a greater area- under-the curve (AUC) per unit done, resulting in a greater time in the therapeutic window per unit dose.
  • AUC area- under-the curve
  • compositions or formulations comprising PEGylated aptamers described herein can comprise an aptamer concentration of from about 20 mg/mL to about 60 mg/mL, from about 35 mg/mL to about 80 mg/mL, or from about 35 mg/mL to about 70 mg/mL.
  • the formulation can comprise an aptamer concentration of at least about 20 mg/mL, at least about 25 mg/mL, at least about 30 mg/mL, at least about 35 mg/mL, at least about 40 mg/mL, at least about 45 mg/mL, at least about 50 mg/mL, or at least about 60 mg/mL.
  • attaching a PEG reagent comprising two PEG arms, three PEG arms, or more to an aptamer may not interfere with the ability of the aptamer to bind its target.
  • attaching a PEG reagent comprising two, three or more PEG arms to an aptamer may improve, may impede, or may have no impact, on the ability of the aptamer to inhibit a function associated with its target.
  • an aptamer attached to a PEG reagent having two or more arms may have substantially the same IC50 for its target in a given assay, as compared to an aptamer that is not attached to a PEG molecule.
  • an aptamer attached to a PEG reagent may have a lower IC50 (j-e., may be more potent) for its target in a given assay, as compared to an aptamer that is not attached to a PEG molecule.
  • the PEGylated aptamers provided herein may often be highly functional.
  • a PEGylated aptamer provide herein has an IC50 of from about 10 pM to about 100 nM.
  • the PEGylated aptamer has an IC50 of at least 10 pM, at least 20 pM, at least 30 pM, at least 40 pM, at least 50 pM, at least 60 pM, at least 70 pM, at least 80 pM, at least 90 pM, at least 100 pM, at least 1 nM, at least 10 nM, at least 20 nM, at least 30 nM, at least 40 nM, at least 40 nM, at least 60 nM, at least 70 nM, at least 80 nM, at least 90 nM, at least 100 nM, or more.
  • the PEGylated aptamer has an IC50 of about 2 nM, about 3 nM, about 3.1 nM, about 4.3 nM, about 4.9 nM, about 5.5 nM, about 5.6 nM, about 7 nM, about 6.3 nM, about 7 nM, or about 10 nM. In some cases, the PEGylated aptamer has an IC50 of less than 10 pM, less than 100 pM, less than 1 nM, less than 20 nM, less than 40 nM, less than 60 nM, less than 80 nM, or less than 100 nM.
  • a PEGylated aptamer provided herein contains PEG reagents with two or more arms, three or more arms, four or more arms, etc., and may have an IC50 of less than about 100 nM, less than about 80 nM, less than about 60 nM, less than about 40 nM, less than about 20 nM, less than about 10 nM, less than about 6 nM, less than about 5 nM, less than about 4 nM, or even lower.
  • the PEG reagents in such aptamers have a molecular weight of about 10 kDa, about 20 kDa, about 30 kDA, about 40 kDa, about 50 kDa, about 60 kDa, about 70 kDa, or about 80 kDa.
  • PEGylated aptamers that are particularly useful in injection devices, such as pre-filled syringes, as they may maintain injectability even when formulated at higher aptamer concentrations (e.g ., greater than 20 mg/ml, greater than 30 mg/ml, or higher).
  • PEGylated aptamers contain PEG reagents with multiple arms or branches (e.g., greater than 3 arms, greater than 4 arms).
  • PEG reagents comprising multiple PEG arms (e.g, 3 or more PEG arms) may change or impact the viscosity of the resulting formulations such that higher aptamer concentrations of PEGylated aptamers may be injected.
  • the PEGylation of aptamers with PEG reagents described herein can result in formulations that are suitable for injection, particularly for intravitreal or intraocular injections. High viscosity may require higher injection forces, making the formulation undesirable for delivery by intravitreal or intraocular injection. As such, the reduction in viscosity described herein may make the disclosed PEGylated aptamers especially useful for intravitreal injection, or other types of injection.
  • Injection break force is the viscous resistive force required to move the plunger of the syringe, measured in Newtons (N), or in other words, the force required to initiate movement of the plunger.
  • Injection break force may refer to the amount of force required to expel a formulation from a needle and may at least partly be a function of the viscosity of the formulation (as well as needle gauge, syringe barrel specifications, etc.).
  • a formulation containing a PEG reagent with a greater number of arms may have decreased injection break force during administration when compared to a formulation that contains the same aptamer attached to a PEG reagent having fewer arms, even when the PEG reagents have comparable molecular weights.
  • a formulation comprising an aptamer attached to a PEG reagent having 3 or more PEG arms has a lower injection break force than a formulation comprising the same aptamer attached to a PEG reagent having 2 or less PEG arms, when tested under the same conditions (e.g, same aptamer concentration of PEGylated aptamer, same needle gauge, etc.).
  • a formulation that comprises a PEGylated aptamer of the disclosure can have an injection force of less than about 15 N, less than about 13 N, less than about 12 N, less than about 10 N, less than about 8 N, less than about 5 N, or less than about 1 N.
  • a formulation that comprises a PEGylated aptamer of the disclosure can have an injection force of at least about 1 N, at least about 2 N, at least about 4 N, at least about 5 N, at least about 10N, or at least about 12 N, or at least about 15 N.
  • a formulation that comprises a PEGylated aptamer of the disclosure can have an injection force of at least about 1 N.
  • a formulation that comprises a PEGylated aptamer of the disclosure can have an injection force of at most about 15 N.
  • a formulation that comprises a PEGylated aptamer of the disclosure can have an injection force of at most about less than 13 N. In some instances, a formulation comprising a PEGylated aptamer of the disclosure can have an injection force of at most about 20 N. In some instances, a formulation that comprises a PEGylated aptamer of the disclosure can have an injection force of about 1 N, about 3 N, about 5 N, about 7 N, about 9 N, about 11 N, about 13 N, or about 15 N.
  • a formulation provided herein may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 30 mg/mL and having an injection force of less than about 20 N through a syringe needle.
  • a formulation may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 35 mg/mL and having an injection force of less than about 20 N.
  • a formulation may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 35mg/mL and having an injection force of less than about 13N.
  • a formulation provided herein may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 40 mg/mL and having an injection force of less than about 13N through a syringe needle. In some cases, a formulation may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 50 mg/mL and having an injection force of less than about 13N. In some cases, a formulation may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 35mg/mL and having an injection force of less than about 25 N.
  • a formulation may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 30 mg/mL and having an injection force of less than about 13N when used with a 28-gauge needle. In some cases, a formulation may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 30 mg/mL and having an injection force of less than about 13N when used with a 27-gauge needle. In some cases, a formulation may comprise a PEGylated aptamer of the disclosure having an aptamer concentration greater than or about 30 mg/mL and having an injection force of less than about 13N when used with a 29 or 30-gauge needle.
  • This disclosure also provides methods of making PEGylated aptamers.
  • this disclosure provides methods of making PEGylated aptamers with reduced viscosity when present at a certain concentration.
  • the method may involve attaching an aptamer to a PEG reagent with multiple arms ( e.g three or more arms, four or more arms, etc.) in order to reduce the viscosity of a formulation containing the PEGylated aptamer.
  • a formulation is administered to the eye of a subject to treat an ocular disease or disorder, for example, macular degeneration, dry AMD, geographic atrophy, wet AMD or Stargardt disease.
  • an ocular disease or disorder for example, macular degeneration, dry AMD, geographic atrophy, wet AMD or Stargardt disease.
  • the ocular disease is a diabetes related eye disease, but is not limited to diabetic retinopathy (DR), peripheral diabetic retinopathy (PDR) or diabetic macular edema (DME).
  • DR diabetic retinopathy
  • PDR peripheral diabetic retinopathy
  • DME diabetic macular edema
  • the ocular disease is a macular edema; macular edema includes, but is not limited to, retinal vein occlusion (RVO).
  • Other ocular diseases and disorders include, but are not limited to, Polyploidal Choroidal
  • PCV Vasculopathy
  • mCNV Myopic Choroidal Neovascularization
  • Vitreoretinopathy Glaucoma, Uveitis, Keratitis, ocular hypertension, a retinoblastoma, Blepharitis, Nystagmus, Retinitis, a sub conjunctive hemorrhage, or a combination thereof.
  • a composition of the disclosure can be administered once or more than once each day.
  • the composition is administered as a single dose (i.e., one-time use).
  • the single dose may have a therapeutic effect and may even be curative.
  • the composition may be administered serially (e.g., administered every day, every week, twice a week, or once a month without a break for the duration of the treatment regimen).
  • the total therapeutically effective treatment regime can occur over a period of time less than a week, less than a month, less than 2 months, less than 3 months, less than 4 months, less than 5 months, or less than 6 months.
  • the composition is administered over a period of at least 4 weeks, at least 8 weeks, at least 12 weeks, at least 16 weeks, at least every 20 weeks, at least every 24 weeks or higher.
  • a therapeutically effective amount can be administered at a frequency of at most once per week, at most once every 4 weeks, at most once every eight weeks, at most once every twelve weeks, at most once every sixteen weeks, at most once every twenty weeks, or at most once every twenty-four weeks or greater.
  • therapeutic compositions provided herein can be administered once a month (approximately 28 days) for about three months, about 4 months, about 5 months or about 6 months, followed by less frequent dosing schedule based on clinical assessment.
  • the less frequent dosing schedule can comprise 4-5 dose administrations over 9 months, or one dose administration every 3 months.
  • Clinical assessment can include monitoring of retinal thickness (e.g ., center point thickness or central foveal thickness), leakage from choroidal neovascularization, microvascular retinal change, neovascularization, or a combination thereof.
  • Other methods of assessing a subjects response to treatment of the various indications are known in the art and are contemplated for use herein.
  • compositions provided herein are especially useful for injections, e.g., intravitreal or intraocular injection.
  • injections e.g., intravitreal or intraocular injection.
  • different types of injections e.g., intravitreal or intraocular injection.
  • a formulation of the disclosure is administered by local ocular delivery.
  • local ocular delivery include intravitreal (IVT), intracamarel, subconjunctival, subtenon, retrobulbar, posterior juxtascleral, and peribulbar.
  • IVT intravitreal
  • Local ocular delivery may generally involve injection of a liquid formulation.
  • a formulation of the disclosure is administered systemically.
  • Systemic administration can involve oral administration.
  • systemic administration can be intravenous administration, subcutaneous
  • formulations suitable for delivery of the pharmaceutical compositions described herein may include a sustained release gel or polymer formulations by surgical implantation of a biodegradable microsize polymer system, e.g, microdevice, microparticle, or sponge, or other slow release transscleral devices, implanted during the treatment of an ophthalmic disease, or by an ocular delivery device, e.g, polymer contact lens sustained delivery device.
  • the formulation is a polymer gel, a self-assembling gel, a durable implant, an eluting implant, a biodegradable matrix or biodegradable polymers.
  • the formulation may be administered by iontophoresis using electric current to drive the composition from the surface to the posterior of the eye.
  • the formulation may be administered by a surgically implanted port with an intravitreal reservoir, an extra-vitreal reservoir or a combination thereof.
  • implantable ocular devices can include, without limitation, the DURASERTTM technology developed by Bausch & Lomb, the ODTx device developed by On Demand Therapeutics, the Port Delivery System developed by FORSIGHT ® VISION4 and the Replenish MICROPUMP ® System developed by Replenish, Inc.
  • nanotechnologies can be used to deliver the pharmaceutical compositions including, but not limited to, nanospheres, nanoparticles, nanocapsules, liposomes, nanomicelles and dendrimers.
  • the vitreous humor (also referred to herein as the“vitreous”) is a jelly-like substance that fills the space between the lens and the retina of the eyeball of humans and other vertebrates. Because the vitreous humor is in direct contact with the retina, intravitreal administration of therapeutic agents (i.e., administration directly to the vitreous humor) may be utilized for the treatment of retinal diseases.
  • IVVT half-life may be used interchangeably and refer to the amount of time required for the amount of a substance (e.g a therapeutic agent) to drop by half the amount in the vitreous. For example, if the concentration of a therapeutic agent injected into the vitreous falls by 50% in 8 days, the therapeutic agent would have an IVT half-life of 8 days.
  • the IVT half-life can be affected by a multitude of factors, non limiting examples including stability of the agent, clearance of the agent from the vitreous, and interactions of the agent with the vitreous environment.
  • compositions that have an improved IVT half-life.
  • the compositions may be therapeutic agents, in some cases therapeutic agents that are used to treat ocular diseases, which are modified or altered to improve the IVT half-life.
  • PEGylation of aptamers with a PEG reagent comprising two, three or more PEG arms may prolong the intravitreal half-life of the PEGylated aptamers.
  • the PEGylated aptamer can have an intravitreal half-life of from about 110 hours to about 180 hours or more in a rabbit.
  • the PEGylated aptamer can have an intravitreal half-life of about 4 days, about 5 days, about 6 days, about 8 days, about 10 days, about 12 days, about 15 days, about 18 days, or about 20 days in a human.
  • Aptamers can be PEGylated with the PEG reagents described herein and maintain high affinity for the intended target of the aptamer.
  • the PEGylated aptamer has a dissociation constant (K d ) value of from about 10 pM to about 100 nM.
  • the PEGylated aptamer has a K d value of at least about 10 pM.
  • the PEGylated aptamer has a K value of at most about 100 nM.
  • the PEGylated aptamer has a K d value of about 10 pM, about 100 pM, about 1 nM, about 5 nM, about 10 nM, about 20 nM, about 40 nM, about 60 nM, or about 80 nM.
  • the PEGylated aptamers described herein can have sufficient metabolic stability to persist in the eye.
  • a PEGylated aptamer can maintain an intravitreal concentration from about 100 mM to about lnM for at least 8 weeks.
  • the aptamer can maintain an intravitreal concentration from about 100 mM to about O.lnM for at least 12 weeks.
  • a PEGylated aptamer can maintain an intravitreal concentration from about 100 pM to about 0. lnM for at least 16 weeks.
  • a PEGylated aptamer can maintain an intravitreal concentration from about 100 pM to about O.lnM for at least 20 weeks.
  • a PEGylated aptamer can maintain an intravitreal concentration from about 100 pM to about O.lnM for at least 24 weeks. In some instances, a PEGylated aptamer can maintain an intravitreal concentration from about 100 pM to about 0. lnM for at least greater than 24 weeks.
  • the ocular disease is macular degeneration.
  • macular degeneration is age-related macular degeneration (AMD).
  • AMD age-related macular degeneration
  • AMD can be dry age-related macular degeneration.
  • Dry age-related macular degeneration can be advanced dry age-related macular degeneration ( i.e geographic atrophy).
  • the ocular disease is juvenile macular degeneration.
  • the ocular disease is wet age- related macular degeneration.
  • the ocular disease is Stargardt disease.
  • the ocular disease is a diabetes related eye disease, but is not limited to diabetic retinopathy (DR), peripheral diabetic retinopathy (PDR) or diabetic macular edema (DME).
  • DR diabetic retinopathy
  • PDR peripheral diabetic retinopathy
  • DME diabetic macular edema
  • the ocular disease is a macular edema; macular edema includes, but is not limited to, retinal vein occlusion (RVO).
  • macular edema includes, but is not limited to, retinal vein occlusion (RVO).
  • Other ocular diseases and disorders include, but are not limited to, Polyploidal Choroidal Vasculopathy (PCV), Myopic Choroidal
  • Neovascularization mCNV
  • Glaucoma Glaucoma
  • Uveitis Uveitis
  • Keratitis ocular hypertension
  • a retinoblastoma Blepharitis
  • Nystagmus a retinoblastoma
  • Retinitis a sub conjunctive hemorrhage, or a combination thereof.
  • the methods and compositions provided herein may alleviate or reduce (partially or completely) one or more symptoms of an ocular disease.
  • the one or more symptoms of the ocular disease are reduced at least 2-fold, at least 3-fold, at least 4- fold, at least 5-fold, at least lO-fold or more compared to when a therapeutic is used that has two or fewer PEG arms.
  • treatment with an aptamer (or oligonucleotide) provided herein may result in a reduction in the severity of symptoms associated with an ocular disease.
  • treatment with an aptamer (or oligonucleotide) described herein may slow, halt or reverse the progression of one or more symptoms associated with ocular disease.
  • treatment with an aptamer (or oligonucleotide) described herein may prevent the development symptoms associated with an ocular disease (i.e., improvement in visual acuity, reduction in central foveal thickness, reduction in intra- and/or subretinal fluid).
  • treatment with an aptamer (or oligonucleotide) described herein may slow, halt or reverse the progression of a disease, as measured by the number and severity of symptoms experienced.
  • the terms“subject” and“patient” may be used interchangeably herein to refer to a vertebrate, preferably a mammal, more preferably a human.
  • the methods provided herein may be used to treat a subject in need thereof with a composition of the disclosure (e.g ., a PEGylated aptamer as described herein).
  • the subject has or is suspected of having an ocular disease or disorder.
  • the subject is a human.
  • the human can be an adult (e.g., older than 18 years old).
  • the subject can be a child (e.g, from birth to 18 years old).
  • the subject can be a child from birth to less than 10 years old.
  • the subject can be a pre-term infant.
  • the subject can be a full-term infant.
  • the subject may be a non-human mammal.
  • Non-human mammals may include, but are not limited to, murine animals, simian animals, laboratory animals, farm animals, sport animals, and pets.
  • Non-limiting examples of a non-human mammal include, but are not limited to, primates such as an ape (e.g, a chimpanzee, a baboon, a gorilla, or an orangutan), an old world monkey (e.g, a rhesus monkey), a new world monkey, or any other non-human primate.
  • non human mammal examples include, but are not limited to, a dog, a cat, a rodent (e.g., a rat, a mouse, a rabbit, etc.), a horse, a cow, an alpaca, a llama, etc.
  • rodent e.g., a rat, a mouse, a rabbit, etc.
  • horse e.g., a cow, an alpaca, a llama, etc.
  • the methods provided herein include administering the compositions of the disclosure to tissues, cells, and their progeny of a biological entity obtained in vivo or cultured in vitro.
  • the methods provided herein include testing the PEGylated aptamer compositions in assays including cell culture assays or in vivo animal models.
  • Such methods may include using the PEGylated aptamers of the disclosure in affinity -binding assays (e.g, to measure 3 ⁇ 4 values), or to test the efficacy of the PEGylated aptamers to inhibit a function associated with a target (e.g, to measure IC 50 values).
  • the assays used herein may be selected based on the aptamer target and the biological pathway to be studied. Also encompassed herein are methods of testing the PEGylated aptamers in animal models to measure pharmacodynamics and/or
  • compositions herein may include any number of pharmaceutical compositions for the treatment of ocular diseases or disorders as well as any type of formulation containing a PEGylated aptamer provided herein.
  • the pharmaceutical compositions may include a therapeutically effective amount of any composition as described herein (e.g ., a therapeutic aptamer conjugated to a PEG reagent).
  • the formulation or pharmaceutical composition provided herein contains a PEGylated aptamer provided herein and another substance or component provided herein, such as a liquid or buffer.
  • the pharmaceutical composition or formulation is solely composed of PEGylated aptamers.
  • the formulation or pharmaceutical composition is substantially composed of PEGylated aptamers (e.g., greater than 70%, greater than 80%, greater than 90%, greater than 95% composed of PEGylated aptamers).
  • the formulation or pharmaceutical composition is mostly composed of PEGylated aptamers (e.g, greater than 50% PEGylated aptamers).
  • the PEGylated aptamer is a minor constituent of the pharmaceutical formulation.
  • the PEGylated aptamer makes up less than 20%, less than 10%, or less than 5% of the pharmaceutical formulation or composition.
  • the PEGylated aptamer makes up from about 3% to about 5% of the pharmaceutical formulation or composition.
  • the formulation or pharmaceutical composition may further include any number of excipients, vehicles or carriers.
  • the pharmaceutical composition may include a therapeutically effective amount of the composition, alone or in combination, with one or more vehicles (e.g, pharmaceutically acceptable compositions or e.g, pharmaceutically acceptable carriers).
  • Excipients may include any and all buffers, solvents, lubricants, preservatives, diluents, and vehicles (or carriers). Generally, the excipient is compatible with the compositions described herein.
  • the pharmaceutical composition may also contain minor amounts of non-toxic auxiliary substances such as wetting or emulsifying agents, pH buffering agents, and other substances such as, for example, sodium acetate, and
  • a therapeutically effective amount of the composition is
  • a therapeutic or desired response refers to an amount of the composition that provokes a therapeutic or desired response in a subject.
  • the therapeutic or desired response is the alleviation or reduction of one or more symptoms associated with a disease or disorder.
  • a therapeutic or desired response is prophylactic treatment of a disease or a disorder.
  • the pharmaceutical compositions may be administered in a dose that is sufficient to cause a therapeutic benefit to or a therapeutic response in the subject.
  • the dose may vary depending on a variety of factors including the aptamer and the PEG reagent selected for use.
  • a therapeutically effective amount of a PEGylated aptamer of the disclosure e.g, an aptamer attached to a PEG having 2, 3 or more arms
  • a therapeutically effective amount of an aptamer attached to a PEG reagent having 2 or more arms may be administered to a subject in a smaller volume than an aptamer attached to a PEG reagent having less than 2 arms.
  • a therapeutically effective amount of an aptamer attached to a PEG reagent having 3 or more arms may be administered to a subject in a smaller volume than an aptamer attached to a PEG reagent having less than 3 arms.
  • a formulation comprising a PEGylated aptamer of the disclosure may be more concentrated (and hence, require a smaller administration volume).
  • the therapeutic composition/formulation may enable a therapeutically effective amount to be delivered to a subject in a single administration, e.g, a single injection, a single intravitreal injection.
  • the therapeutic composition/formulation may possess a viscosity that enables a therapeutically effective amount to be delivered to a subject in a single administration, e.g, a single injection, a single intravitreal injection.
  • a therapeutically effective amount of an aptamer attached to a PEG reagent having 3 or more arms may be less than a therapeutically effective amount of an aptamer attached to a PEG reagent having two or less arms. Without wishing to be bound by theory, this may be because an increased intravitreal retention time may reduce the amount of PEGylated aptamer needed to achieve a therapeutic response.
  • the pharmaceutical compositions herein generally may be administered by injection to the vitreous (i.e., intravitreal (IVT) administration).
  • IVT administration may be to one eye if only one eye is affected by the ocular disease, or to both eyes if both eyes are affected.
  • the pharmaceutical compositions herein may be in a formulation suitable for intravitreal administration.
  • the pharmaceutical compositions may be prepared in a liquid formulation for injection into the vitreous.
  • Liquid formulations provided herein may have low viscosity, e.g., a viscosity amenable to intravitreal injection, yet may also contain a relatively high concentration of PEGylated aptamer (e.g, about 25 mg/mL to about 60 mg/mL as calculated on the molecular weight of the aptamer moiety only).
  • a relatively high concentration of PEGylated aptamer e.g, about 25 mg/mL to about 60 mg/mL as calculated on the molecular weight of the aptamer moiety only.
  • the pharmaceutical composition may comprise a PEGylated aptamer concentration of at least about 25 mg/mL, at least about 30 mg/mL, at least 35 mg/mL, at least 40 mg/mL, at least 45 mg/mL, at least 50 mg/mL or at least 60 mg/mL as calculated on the molecular weight of the aptamer moiety only.
  • a liquid formulation provided herein may have an aptamer concentration of PEGylated aptamer of greater than about 25 mg/ml or greater than about 30 mg/ml when formulated for intravitreal administration.
  • a liquid formulation provided herein may have an aptamer concentration of PEGylated aptamer of greater than about 35 mg/ml when formulated for intravitreal administration. In another specific example, a liquid formulation provided herein may have an aptamer concentration of PEGylated aptamer of greater than about 40 mg/ml when formulated for intravitreal administration.
  • the pharmaceutical composition may comprise a PEGylated aptamer concentration of at least about 25 mg/mL, at least about 30 mg/mL, at least 35 mg/mL, at least 40 mg/mL, at least 45 mg/mL, at least 50 mg/mL or at least 60 mg/mL, wherein the composition has a viscosity as described in Table 2.
  • the pharmaceutical composition may comprise a PEGylated aptamer concentration of at least about 25 mg/mL, at least about 30 mg/mL, at least 35 mg/mL, at least 40 mg/mL, at least 45 mg/mL, at least 50 mg/mL or at least 60 mg/mL, wherein the composition has a viscosity as described in Table 3.
  • the pharmaceutical composition may comprise a PEGylated aptamer concentration of at least about 25 mg/mL, at least about 30 mg/mL, at least 35 mg/mL, at least 40 mg/mL, at least 45 mg/mL, at least 50 mg/mL or at least 60 mg/mL, wherein the composition has a viscosity between 5 and 90.0 centipoise (cP).
  • a PEGylated aptamer concentration of at least about 25 mg/mL, at least about 30 mg/mL, at least 35 mg/mL, at least 40 mg/mL, at least 45 mg/mL, at least 50 mg/mL or at least 60 mg/mL, wherein the composition has a viscosity between 5 and 90.0 centipoise (cP).
  • the pharmaceutical composition may comprise a PEGylated aptamer concentration of at least about 25 mg/mL, at least about 30 mg/mL, at least 35 mg/mL, at least 40 mg/mL, at least 45 mg/mL, at least 50 mg/mL or at least 60 mg/mL, wherein the composition has a viscosity between 1 and 260 cP.
  • the pharmaceutical composition may comprise a PEGylated aptamer concentration of at least about 25 mg/mL, at least about 30 mg/mL, at least 35 mg/mL, at least 40 mg/mL, at least 45 mg/mL, at least 50 mg/mL or at least 60 mg/mL, wherein the composition has a viscosity less than 300 cP, less than 250 cP, less than 200 cP, less than 150 cP, less than 100 cP, less than 90 cP, less than 80 cP, less than 70 cP, less than 60 cP, less than 50 cP, less than 40 cP, less than 30 cP, less than 20 cP, less than 10 cP, or less than 5 cP.
  • a viscosity less than 300 cP, less than 250 cP, less than 200 cP, less than 150 cP, less than 100 cP, less than 90 cP, less than 80 cP, less than 70
  • This disclosure provides multi-armed (e.g., 4-armed) PEGylated aptamers with reduced viscosity at higher concentrations of aptamer.
  • the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 200 cP.
  • the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 100 cP.
  • the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 30 cP.
  • the concentration of the aptamer is greater than 35 mg/ml and the viscosity is less than 200 cP.
  • the concentration of the aptamer is greater than 35 mg/ml and the viscosity is less than 100 cP. In some cases, the concentration of the aptamer is greater than 25 mg/ml and the viscosity is less than 200 cP. In some cases, the concentration of the aptamer is greater than 25 mg/ml and the viscosity is less than 30 cP.
  • the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 200 cP. In some cases, the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 100 cP. In some cases, the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 30 cP.
  • the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 200 cP. In some cases, the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 100 cP.
  • the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 30 cP. In some cases, the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 35 mg/ml and the viscosity is less than 200 cP.
  • the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 35 mg/ml and the viscosity is less than 100 cP. In some cases, the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 25 mg/ml and the viscosity is less than 200 cp. In some cases, the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 25 mg/ml and the viscosity is less than 30 cp.
  • the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 200 cP. In some cases, the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 100 cP. In some cases, the aptamer is at least 40 kDa and the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 30 cP.
  • the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 200 cP. In some cases, the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 100 cP.
  • the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 30 mg/ml and the viscosity is less than 30 cP. In some cases, the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 35 mg/ml and the viscosity is less than 200 cP.
  • the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 35 mg/ml and the viscosity is less than 100 cP. In some cases, the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 25 mg/ml and the viscosity is less than 200 cP. In some cases, the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 25 mg/ml and the viscosity is less than 30 cP.
  • the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 200 cP. In some cases, the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 100 cP. In some cases, the aptamer is at least 60 kDa and the concentration of the aptamer is greater than 45 mg/ml and the viscosity is less than 30 cP.
  • a liquid formulation as provided herein may be formulated in a pre- filled syringe.
  • a liquid formulation may be formulated in a volume of 10 pL, 20 pL, 30 pL, 40 pL, 50 pL, 60 pL, 70 pL, 80 pL, 90 pL, 100 pL or greater than 100 pL.
  • pre-filled syringes that contain a composition that comprises any of the PEGylated aptamers described herein.
  • PEGs suitable for intravitreal administration of a PEG-aptamer at a formulation strength of 30 mg/mL using a 1 ⁇ 2 inch 30 G needle may be linear PEGs less than or equal to approximately 40 kDa, or less than approximately 60 kDa; Y-armed PEGs less than or equal to 60 kDa; multi-arm PEGs with an architecture per PEG Reagent 2 of less than or equal to 60 kDa; and multi-arm PEGs with an architecture per PEG Reagent 3 of less than or equal to 40 kDa, less than or equal to 60 kDa, or less than or equal to 80 kDa.
  • PEGs suitable for intravitreal administration of a PEG-aptamer at a formulation strength of 30 mg/mL using a 1 ⁇ 2 inch 29 G needle may be linear PEGs less than or equal to approximately 40 kE)a or less than approximately 60 kE)a; Y-armed PEGs less than or equal to 60 kDa; multi-arm PEGs with an architecture per PEG Reagent 2 of less than or equal to 60 kE)a; and multi-arm PEGs with an architecture per PEG Reagent 3 of less than or equal to 40 kE)a, less than or equal to 60 kE)a, or less than or equal to 80 kE)a.
  • PEGs suitable for intravitreal administration of a PEG-aptamer at a formulation strength of 50 mg/mL using a 1 ⁇ 2 inch 30 G needle may be linear PEGs less than or equal to approximately 40 kDa or less than approximately 60 kDa; Y-armed PEGs less than or equal to 40 kDa; multi-arm PEGs with an architecture per PEG Reagent 2 of less than or equal to 60 kDa; and multi-arm PEGs with an architecture per PEG Reagent 3 of less than or equal to 40 kDa, less than or equal to 60 kDa, or less than or equal to 80 kDa.
  • PEGs suitable for intravitreal administration of a PEG-aptamer at a formulation strength of 50 mg/mL using a 1 ⁇ 2 inch 29 G needle may be linear PEGs less than or equal to approximately 40 kDa or less than approximately 60 kDa; Y-armed PEGs less than or equal to 60 kDa; multi-arm PEGs with an architecture per PEG Reagent 2 of less than or equal to 60 kDa; and multi-arm PEGs with an architecture per PEG Reagent 3 of less than or equal to 40 kDa, less than or equal to 60 kDa, or less than or equal to 80 kDa.
  • polydispersity index refers to a measure of the distribution of molecular mass in a given polymer sample.
  • the polydispersity index therefore, reflects the level of uniformity in a sample.
  • the therapeutic compositions provided herein may have a PDI of less than 1.05.
  • the molecular mass of PEGylated aptamers present in a therapeutic composition of the disclosure may be relatively uniform.
  • the PDI of a therapeutic compositions may be less than 1.05, less than 1.04, less than 1.03, less than 1.02, less than 1.01, or about 1 00
  • compositions described herein may be co-administered with one or more additional therapeutic agents.
  • the one or more additional therapeutic agents may be conjugated to a PEG reagent as described herein or may be unconjugated.
  • the one or more additional therapeutic agents enhance or act synergistically in combination with the compositions provided herein.
  • the PEGylated aptamer may be administered to a subject by ocular delivery. In some instances, the PEGylated aptamer is administered by intravitreal injection. In other cases, the PEGylated aptamer is administered by periocular injection.
  • Example 1 Viscosity and Intravitreal Half-lives of three PEG Reagents attached to an Identical Aptamer
  • FIG. 1 provides three PEG reagents of the same molecular weight of about 40kDa but with different branching configurations.
  • the respective PEGs shown in FIG.l were conjugated to Aptamer 15 (see Table 1), as follows. Briefly, a concentrated feed solution consisting of aptamer in DMSO, 16 to 25 mM borate and water was combined with a solution consisting of several equivalents of the respective PEG reagents shown in FIG.
  • each PEG-aptamer was purified by anion exchange chromatography to collect the pegylated aptamer and remove unreacted PEG and unreacted aptamer.
  • Anion exchange purified PEG-aptamers were desalted by ultrafiltration into water prior to functional characterization.
  • the pegylated version of Aptamer 15 attached to the respective PEGs are Aptamer P01, Aptamer P02 and Aptamer P03. respectively.
  • Each sample was prepared at a target concentration of 210 mg/mL, 168 mg/mL, 105 mg/mL, and 42 mg/mL (total weight including PEG and aptamer). Each prepared sample was incubated at 80 °C for 10 minutes with the vial cap on and allowed to cool to room temperature for 1 hour. Viscosity testing was performed on each solution using a Brookfield RVDV-II+ cone and plate viscometer utilizing spindle CPE-40 and a sample size of 0.5 mL at 25 °C. A spindle and RPM combination was employed such that a valid torque measurement was obtained for all measurements (> 10% torque). Results are provided in Table 2.
  • the maximal volume of an intravitreal injection to the human eye is limited to less than or equal to approximately 100 uL. Therefore, reducing the viscosity of PEG-aptamer formulations for intravitreal injection to the eye is critical, as PEG-aptamers with reduced viscosity vs. concentration properties can be formulated at higher strength to enable administration of more drug to the eye.
  • the eye is a fragile tissue, which limits both the size of the needle that can be used as well as the injection, or break force, applied to the plunger for intravitreal administration of drugs to the eye. Break force is the viscous resistive force required to move the plunger of the syringe, measured in Newtons (N), or in other words, the force required to initiate movement of the plunger.
  • N Newtons
  • the total injection break force on the plunger is equal to the sum of the viscous resistive force and the friction related to moving the piston. Since in most syringes, there is technology to mitigate this friction, one can assume that this force is negligible when compared to the viscous resistive force.
  • Injection Force Viscous resistive force + Plunger force of friction With the plunger friction component reduced to zero, the injection force (break force) is equal to the viscous resistive force which is calculated from equation 1. Equation 1 can easily be modeled once a syringe/needle combination is selected. With that combination, the injection force (break force) can be estimated for a wide variety of materials for which viscosity data exists.
  • Aptamer P01 comprising PEG Reagent 1 with 2 PEG arms of approximately 20 kDa each, can support a maximum aptamer concentration of about 30 mg/mL when injected into the eye using a 30-gauge needle. Above this concentration, the viscosity of the composition may prevent delivery of the composition by intravitreal injection when a 30-gauge needle is used. As the branching of the PEG reagent increases, the maximum aptamer concentration also increases while maintaining injectability using a high gauge needle.
  • Aptamer P02 comprising PEG Reagent 2, which has four arms of approximately 7.5 kDa each that branch off from two PEG arms of approximately 5 kE)a each, can support an aptamer concentration of about 40 mg/mL when administered to the eye via intravitreal injection using a 30-gauge needle.
  • Aptamer P03 comprising PEG Reagent 3, which has four arms of approximately 10 kE)a each, branching from a linear carbon alcohol backbone, can support an aptamer concentration greater than 50 mg/mL when administered to the eye via an intravitreal injection using a 30-gauge needle.
  • the branching not only affects the maximum aptamer concentration that can be achieved while maintaining injectability, but also the intravitreal half-life.
  • Aptamer P01 comprising PEG Reagent 1 containing 2 PEG arms had an intravitreal half- life of about 110 hours in rabbits.
  • PEG Reagent 2 Aptamer P02
  • PEG Reagent 3 Aptamer P03
  • the intravitreal half-life increased to 130 hours. This indicated that increasing the number of PEG arms of a PEG reagent without changing the total molecular weight increased the intravitreal half-life of a PEGylated aptamer, making it potentially more effective as a therapeutic composition.
  • Therapeutic PEGylated aptamers that can be administered at greater doses and/or with relatively longer intravitreal half-life can be more desirable as a therapy because they would result in fewer injections for the delivery of the desired effective amount. Fewer injections can improve safety by potentially reducing the risk of injury and/or infection.
  • therapeutic PEGylated aptamers with relatively lower viscosity can be injected intravitreally using higher gauge needles (i.e., smaller needle bore sizes) which can have several benefits including reducing the risk of injury to the patient and enhancing patient comfort.
  • Example 2 Polyethylene glycol carriers suitable for administration of PEG- aptamer by intravitreal administration
  • the PEG moiety of the aptamer must not contribute a viscosity level greater than required for intravitreal administration in a suitable syringe/needle format.
  • the viscosity vs. concentration profile of a range of PEGs with varying molecular weights and architectures was determined using the methods provided in Example 1. Once the viscosity at a given concentration is determined, the injection break force for a given syringe and needle configuration can be calculated as described in Example 1.
  • the viscosity of a PEG-aptamer formulation is dependent on the viscosity versus concentration profile of the aptamer moiety, the PEG moiety and the PEG-aptamer moiety.
  • the maximum suitable formulation viscosity for a 1 ⁇ 2 inch 30G needle may be less than or equal to 90 cP, and for a 1 ⁇ 2 inch 29G needle may be less than approximately 145 cP, assuming an injection break force limit of less than 13N.
  • PEGs suitable for intravitreal administration will provide for a viscosity versus concentration profile such that the viscosity of the PEG at any given concentration, normalized for a given PEG-aptamer dose based on the molecular weight of the aptamer moiety only, must be less than that suitable for IVT administration of the PEG-aptamer.
  • Table 3 The data from this analysis is provided in Table 3.
  • PEGs suitable for intravitreal administration of a PEG-aptamer at a formulation strength of 30 mg/mL using a 1 ⁇ 2 inch 30 G needle are linear PEGs less than or equal to approximately 40 kDa, or less than approximately 60 kDa; Y-armed PEGs less than or equal to 60 kDa; multi- arm PEGs with an architecture per PEG Reagent 2 of less than or equal to 60 kDa; and multi- arm PEGs with an architecture per PEG Reagent 3 of less than or equal to 40 kDa, less than or equal to 60 kDa, or less than or equal to 80 kDa.
  • PEGs suitable for intravitreal administration of a PEG-aptamer at a formulation strength of 30 mg/mL using a 1 ⁇ 2 inch 29 G needle are linear PEGs less than or equal to approximately 40 kDa or less than approximately 60 kDa; Y-armed PEGs less than or equal to 60 kDa; multi- arm PEGs with an architecture per PEG Reagent 2 of less than or equal to 60 kDa; and multi- arm PEGs with an architecture per PEG Reagent 3 of less than or equal to 40 kDa, less than or equal to 60 kDa, or less than or equal to 80 kDa.
  • PEGs suitable for intravitreal administration of a PEG-aptamer at a formulation strength of 50 mg/mL using a 1 ⁇ 2 inch 30 G needle are linear PEGs less than or equal to approximately 40 kDa or less than approximately 60 kDa; Y-armed PEGs less than or equal to 40 kDa, multi- arm PEGs with an architecture per PEG Reagent 2 of less than or equal to 60 kDa; and multi- arm PEGs with an architecture per PEG Reagent 3 of less than or equal to 40 kDa, less than or equal to 60 kDa, or less than or equal to 80 kDa.
  • PEGs suitable for intravitreal administration of a PEG-aptamer at a formulation strength of 50 mg/mL using a 1 ⁇ 2 inch 29 G needle are linear PEGs less than or equal to approximately 40 kDa or less than approximately 60 kDa; Y-armed PEGs less than or equal to 60 kDa; multi-arm PEGs with an architecture per PEG Reagent 2 of less than or equal to 60 kDa, and multi-arm PEGs with an architecture per PEG Reagent 3 of less than or equal to 40 kDa; less than or equal to 60 kDa, or less than or equal to 80 kDa.
  • Example 3 Treatment of Geographic Atrophy
  • a therapeutic composition comprising an aptamer attached to a PEG reagent to form a PEGylated aptamer.
  • a PEGylated aptamer has the characteristics described herein.
  • the aptamer can comprise less than or equal to about 30 nucleotides, about 35 nucleotides or about 40 nucleotides.
  • the PEGylated aptamer can have an IC50 about equal to that of an anti- Factor D antigen-binding fragment; an IC50 that is 1.5-fold greater than that of an anti- Factor D antigen-binding fragment, or an IC50 that is less than or equal to 3-fold the IC 50 of an anti- Factor D antigen-binding fragment, as measured by a 10% serum RBC lysis assay.
  • the affinity of the PEGylated aptamer can be from about 0.5 to about 2 nM, about 5 nM or about lOnM.
  • the PEGylated aptamer can be specific to Factor D, with a specificity that is about IO,OOOc, about l,000x or about lOOx that of the PEGylated aptamer’ s specificity to other serine proteases.
  • the PEGylated aptamer can be metabolically stable in the vitreous, with a metabolic stability of at least 90% that of pegaptanib equivalent to that of pegaptanib or greater than that of pegaptanib in a rabbit model.
  • Pegaptanib is a selective vascular endothelial growth factor (VEGF) antagonist.
  • the intravitreal half-life, as measured in a rabbit model can be about 70-80 hours, or 80-90 hours.
  • the intravitreal half-life may be greater than the intravitreal half-life of pegaptanib in the same animal model.
  • the PEGylated aptamer can be formulated with an aptamer concentration of about 30 mg/mL, that can be injected using a 29-, 30-, or 31 -gauge needle and can maintain an aptamer concentration greater than or equal to the IC90 concentration for greater than or equal to 8 weeks.
  • a patient with Neovascular (Wet) Age-Related Macular Degeneration may be treated with a therapeutic PEGylated aptamer composition described herein.
  • the composition may comprise a PEG reagent comprising 2 or more PEG arms, attached to an aptamer of about 40 or less nucleotides.
  • the aptamer may modulate the biological activity of human vascular endothelial growth factor (VEGF).
  • VEGF vascular endothelial growth factor
  • the aptamer may have an affinity of about 0.5 nM to about 10 nM.
  • the composition may comprise an aptamer concentration of about 20 mg/mL or greater.
  • the patient may be treated with three monthly doses followed by less frequent dosing with regular assessment. The less frequent dosing may comprise 4-5 doses on average over a 9 month period.
  • the composition may be delivered to the patient via intravitreal injection using a sterile prefilled syringe and a 28-gauge or 30-gauge injection needle.
  • Intraocular pressure, perfusion of the optic nerve, or both, may be monitored following intravitreal injection.

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Abstract

La présente invention concerne des aptamères PEGylés qui contiennent des réactifs PEG présentant des configurations de ramification utiles pour des applications thérapeutiques, telles qu'une injection intraoculaire. La présente invention concerne également des méthodes de traitement de troubles oculaires impliquant l'administration d'aptamères PEGylés thérapeutiques présentant des configurations de ramification à un sujet atteint d'un trouble oculaire.
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US11786510B2 (en) 2021-04-30 2023-10-17 Perfuse Therapeutics, Inc. Pharmaceutical compositions and intravitreal drug delivery systems for the treatment of ocular diseases
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WO2021087399A1 (fr) * 2019-10-30 2021-05-06 Perfuse Therapeutics, Inc. Traitement de maladies oculaires à l'aide d'antagonistes du récepteur de l'endothéline
US11738007B2 (en) 2019-10-30 2023-08-29 Perfuse Therapeutics, Inc. Treatment of glaucoma using endothelin receptor antagonists
US11873279B2 (en) 2020-02-06 2024-01-16 Perfuse Therapeutics, Inc. Compositions for treatment of ocular diseases
US11786510B2 (en) 2021-04-30 2023-10-17 Perfuse Therapeutics, Inc. Pharmaceutical compositions and intravitreal drug delivery systems for the treatment of ocular diseases

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