WO2024173582A2 - Formulations d'oligomères antisens - Google Patents

Formulations d'oligomères antisens Download PDF

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Publication number
WO2024173582A2
WO2024173582A2 PCT/US2024/015838 US2024015838W WO2024173582A2 WO 2024173582 A2 WO2024173582 A2 WO 2024173582A2 US 2024015838 W US2024015838 W US 2024015838W WO 2024173582 A2 WO2024173582 A2 WO 2024173582A2
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Prior art keywords
pharmaceutical formulation
aso
liquid composition
kit
concentration
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PCT/US2024/015838
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English (en)
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WO2024173582A3 (fr
Inventor
Paul Hatala
Surya LAMICHHANE
Tong YUN
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Stoke Therapeutics, Inc.
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Publication of WO2024173582A2 publication Critical patent/WO2024173582A2/fr
Publication of WO2024173582A3 publication Critical patent/WO2024173582A3/fr

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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
    • A61K31/712—Nucleic acids or oligonucleotides having modified sugars, i.e. other than ribose or 2'-deoxyribose
    • 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

Definitions

  • Mutations in the SCN1A gene which is part of the SCN1A-SCN2A-SCN3A gene cluster that encodes alpha-pore forming subunits of the neuronal voltage gated sodium channel, can result in expression of Na V 1.1 protein (also termed as “Na V 1.1”) with reduced functions as compared to a wild- type Na V 1.1 protein, reduced expression of Na V 1.1, or both.
  • Mutations in SCN1A gene are associated with development of numerous diseases and conditions, such as Dravet Syndrome (DS) (Miller, et al., 1993-2015, Gene Reviews, Eds. Pagon RA, et al.
  • suitable dosages and dosage regimens may be discovered through a clinical trial that forms part of the approval process, requires substantial input of intellectual and financial resources of different parties, and is not within the routine of a medical practitioner.
  • patient compliance can be crucial for optimal treatment of various conditions. The more doses required or the more difficult the treatment plan, the less likely a patient is to comply.
  • medical agents have the ability to enhance patients’ quality of life (QOL), they can only do so if they are used correctly.
  • QOL quality of life
  • Suitable pharmaceutical formulations, dosages, dosing regimens and patient populations for treating or reducing the likelihood of developing a disease or condition characterized by a reduced expression or function of NaV1.1 protein in a human subject in need thereof.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: i) an antisense oligomer (ASO), and ii) a pharmaceutically acceptable diluent; wherein the liquid composition is not buffered by a buffering agent; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • ASO antisense oligomer
  • a pharmaceutically acceptable diluent wherein the liquid composition is not buffered by a buffering agent; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: (i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent, wherein (a) the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at -20 °C or a temperature and relative humidity for a time period; (b) the pharmaceutical formulation has a shelf life of a time period when stored at -20 °C or a temperature and relative humidity; (c) the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at - 20 °C or a temperature and relative humidity for a time period; (d) the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at -20 °C or a temperature and relative humidity for a time period; (e) the percent of any unspecified impurities in the pharmaceutical formulation
  • the temperature is 4 °C, 25 °C, 30 °C, 37 °C, or 40 °C.
  • the relative humidity is about 55%, 60%, 65%, 70% or 75%.
  • the time period is at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion
  • the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or
  • each nucleobase of the ASO comprises a modified sugar moiety.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising:(i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent; wherein the liquid composition lacks Na2HPO4 and/or NaH2PO4; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: (i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent consisting of: a) NaCl, b) KCl, c) MgCl 2 or MgCl 2 6H 2 O, and d) CaCl 2 or CaCl 2 2H 2 O.
  • the liquid composition comprises 0.1-50 mM CaCl 2 or CaCl 2 2H 2 O.
  • the liquid composition comprises 1-2 mM CaCl 2 or CaCl 2 2H 2 O. [0015] In some embodiments, the liquid composition comprises about 1.4 mM CaCl 2 or CaCl 2 2H 2 O. [0016] In some embodiments, the liquid composition comprises 0.1-50 mM MgCl 2 or MgCl 2 6H 2 O. [0017] In some embodiments, the liquid composition comprises 0.5-1.5 mM MgCl 2 or MgCl 2 6H 2 O. [0018] In some embodiments, the liquid composition comprises about 0.79 mM MgCl 2 or MgCl 2 6H 2 O.
  • the liquid composition comprises 5-250 mM NaCl, 0.1-20 mM KCl, 0.1- 50 mM CaCl 2 or CaCl 2 2H 2 O, and 0.1-50 mM MgCl 2 or MgCl 2 6H 2 O.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: (i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent; wherein the liquid composition lacks calcium ions and/or magnesium ions; and wherein: (a) the liquid composition comprises potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • the liquid composition comprises a buffering agent.
  • the buffering agent has a pKa at 25 °C of about 4.75; about 5.64; about 1.70, about 6.04 and about 9.09; about 3.1, about 4.7 and about 6.4; or about 6.50.
  • the buffering agent is an effective buffer at a pH range of from about 3.6 to 5.6, from about 5.5 to 6.5, from about 5.5 to 7.4, from about 3.0 to 6.2, or from about 5.8 to 7.2.
  • the buffering agent is selected from the group consisting of acetate, succinate, histidine, citrate, 2-[Bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol (Bis- Tris), and any combination thereof.
  • the liquid composition is formulated for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject.
  • the liquid composition is formulated for administration into cerebrospinal fluid in a brain of a human subject.
  • the ASO comprises at least one modified sugar moiety.
  • each nucleotide of the antisense oligomer comprises a modified sugar moiety.
  • a kit comprising: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; and wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO, and wherein the liquid composition is not buffered by a buffering agent; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • the kit comprises: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; and wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO, and wherein (a) the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at -20 °C or a temperature and relative humidity for a time period; (b) the pharmaceutical formulation has a shelf life of a time period when stored at -20 °C or a temperature and relative humidity; (c) the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at -20 °C or a temperature and relative humidity for a time period; (d)the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of
  • ASO antisense
  • the temperature is 4 °C, 25 °C, 30 °C, 37 °C, or 40 °C.
  • the relative humidity is about 55%, 60%, 65%, 70% or 75%.
  • the time period is at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion
  • the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • the kit comprises: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; and wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO, wherein the liquid composition lacks Na 2 HPO 4 and/or NaH 2 PO 4 ; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • ASO antisense oligomer
  • a pharmaceutically acceptable diluent wherein the concentrate is miscible with the pharmaceutically acceptable diluent
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • the kit comprises: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent consisting of: a) NaCl, b) KCl, c) MgCl 2 or MgCl 2 6H 2 O, and d) CaCl 2 or CaCl 2 2H 2 O, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; and wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO.
  • ASO antisense oligomer
  • the liquid composition comprises 0.1-50 mM CaCl 2 or CaCl 2 2H 2 O. [0037] In some embodiments, the liquid composition comprises 1-2 mM CaCl 2 or CaCl 2 2H 2 O. [0038] In some embodiments, the liquid composition comprises about 1.4 mM CaCl 2 or CaCl 2 2H 2 O. [0039] In some embodiments, the liquid composition comprises 0.1-50 mM MgCl 2 or MgCl 2 6H 2 O. [0040] In some embodiments, the liquid composition comprises 0.5-1.5 mM MgCl 2 or MgCl 2 6H 2 O.
  • the liquid composition comprises about 0.79 mM MgCl 2 or MgCl 2 6H 2 O. [0042] In some embodiments, the liquid composition comprises 5-250 mM NaCl, 0.1-20 mM KCl, 0.1- 50 mM CaCl2 or CaCl22H2O, and 0.1-50 mM MgCl2 or MgCl26H2O.
  • kits comprising: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO, wherein the liquid composition lacks calcium ions and/or magnesium ions; and wherein: (a) the liquid composition comprises potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, WSGR Docket No.47991-744.601 cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • ASO antisense oligomer
  • a pharmaceutically acceptable diluent wherein the concentrate is miscible with the pharmaceutically acceptable diluent
  • mixing the ASO with the pharmaceutically acceptable diluent results in
  • the liquid composition comprises a buffering agent.
  • the buffering agent has a pKa at 25 °C of about 4.75; about 5.64; about 1.70, about 6.04 and about 9.09; about 3.1, about 4.7 and about 6.4; or about 6.50.
  • the buffering agent is an effective buffer at a pH range of from about 3.6 to 5.6, from about 5.5 to 6.5, from about 5.5 to 7.4, from about 3.0 to 6.2, or from about 5.8 to 7.2.
  • the buffering agent is selected from the group consisting of acetate, succinate, histidine, citrate, 2-[Bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol (Bis- Tris), and any combination thereof.
  • the liquid composition is formulated for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject.
  • the liquid composition is formulated for administration into cerebrospinal fluid in a brain of a human subject.
  • the ASO comprises at least one modified sugar moiety.
  • each nucleotide of the antisense oligomer comprises a modified sugar moiety.
  • the kit further comprises: (iii) instructions for diluting or solubilizing the ASO in the pharmaceutically acceptable diluent.
  • the liquid composition comprises 25-250 mM NaCl.
  • the liquid composition comprises 0.1-20 mM KCl.
  • the liquid composition comprises 2-4 mM KCl.
  • the liquid composition about 3 mM KCl.
  • the liquid composition comprises 100-160 mM NaCl.
  • the liquid composition comprises 125-145 mM NaCl. [0059] In some embodiments, the liquid composition comprises about 130 mM NaCl. [0060] In some embodiments, the liquid composition comprises a buffered (pH 6.6 – 7.6) solution. [0061] In some embodiments, the liquid composition comprises 0.1-50 mM Na 2 HPO 4 . [0062] In some embodiments, the liquid composition comprises 0.1-50 mM NaH 2 PO 4 . [0063] In some embodiments, the liquid composition comprises 5-250 mM NaCl, 0.1-20 mM KCl, 0.1- 50 mM Na2HPO4, and 0.1-50 mM NaH2PO4.
  • the ASO is solubilized in the pharmaceutically acceptable diluent.
  • the pharmaceutically acceptable diluent is an isotonic solution.
  • the ASO is not substantially polydisperse.
  • the liquid composition has an osmolality of less than 150 mM.
  • the liquid composition has an osmolality of about 130 mM.
  • the liquid composition further comprises a carbohydrate.
  • the carbohydrate comprises D-glucose.
  • the liquid composition further comprises 1-100 mM D-glucose.
  • the liquid composition further comprises an antioxidant.
  • the antioxidant is t-butylhydroxyquinoline (TBHQ), butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), vitamin E, or any combination thereof.
  • the liquid composition does not comprise a preservative.
  • the liquid composition is packaged in a single-use vial.
  • the liquid composition is formulated or suitable for administration (i) as a bolus injection; (ii) by infusion with a delivery pump; (iii) by intracerebroventricular injection; and/or (iv) by intrathecal injection.
  • the ASO comprises T-methoxyethyl sugar moiety.
  • the T-methoxyethyl sugar moiety is a T-2’-methoxyethyl sugar moiety.
  • the ASO comprises a 2’-O-methoxyethyl moiety.
  • the ASO comprises a thymidine comprising a 2’-O-methoxyethyl moiety.
  • each nucleobase of the ASO comprises a 2’-O-methoxyethyl moiety.
  • the ASO consists of from 8 to 50 nucleobases.
  • the ASO consists of less than 35 nucleobases.
  • the ASO consists of from 16 to 20 nucleobases.
  • the ASO consists of from 12 to 20 nucleobases.
  • the ASO consists of from 8 to 20 nucleobases. [0087] In some embodiments, the ASO comprises a 5’-methylcytosine (5’-MeC). [0088] In some embodiments, each cytosine of the ASO is a 5’-methylcytosine (5’-MeC). [0089] In some embodiments, the ASO comprises a phosphorothioate linkage. [0090] In some embodiments, each internucleoside linkage of the ASO is a phosphorothioate linkage. [0091] In some embodiments, the ASO comprises a locked nucleic acid (LNA).
  • LNA locked nucleic acid
  • the liquid composition comprises 1 mL to 20 mL of the pharmaceutically acceptable diluent, 2 mL to 10 mL of the pharmaceutically acceptable diluent, or 1 mL to 5 mL of the pharmaceutically acceptable diluent.
  • the liquid composition comprises from 0.1 mL to 50 mL of the pharmaceutically acceptable diluent.
  • the liquid composition comprises about 0.1, 0.5, 1, 2, 2.5, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45 or 50 mL of the pharmaceutically acceptable diluent.
  • the liquid composition comprises from about 0.5 milligrams to about 500 milligrams of the ASO.
  • the liquid composition comprises 0.1, 0.5, 1, 2.5, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 22.5, 25, 27.5, 30, 32.5, 35, 37.5, 40, 42.5, 45, 47.5, 50, 52.5, 55, 57.5, 60, 62.5, 65, 67.5, 70, 72.5, 75, 77.5, 80, 82.5, 85, 87.5, 90, 92.5, 95, 97.5, 100, 102.5, 105, 107.5, 110, 112.5, 115, 117.5, 120, 122.5, 125, 127.5, 130, 132.5, 135, 137.5, 140, 142.5, 145, 147.5, 150, 152.5, WSGR Docket No.47991-744.601 155, 157.5, 160, 162.5, 165, 167.5, 170, 172.5, 175, 177.5, 180, 182.5,
  • the ASO is present in the liquid composition at a concentration of from 0.1-500 mg/mL. [0098] In some embodiments, the ASO is present in the liquid composition at a concentration of from 0.1 mg/mL to 250 mg/mL. [0099] In some embodiments, the ASO is present in the pharmaceutical formulation at a concentration of from 6.7 mg/mL to 188 mg/mL or from 6.8 mg/mL to 187 mg/mL.
  • the ASO is present in the liquid composition at a concentration of about 0.1 mg/mL, 0.5 mg/mL, 1 mg/mL, 2 mg/mL, 2.5 mg/mL, 3 mg/mL, 4 mg/mL, 5 mg/mL, 6 mg/mL, 7 mg/mL, 8 mg/mL, 9 mg/mL, 10 mg/mL, 11 mg/mL, 12 mg/mL, 13 mg/mL, 14 mg/mL, 15 mg/mL, 16 mg/mL, 17 mg/mL, 18 mg/mL, 19 mg/mL, or 20 mg/mL.
  • the ASO is present in the liquid composition at a concentration of 11 mg/mL, 22 mg/mL, 33 mg/mL, 44 mg/mL, 55 mg/mL, 66 mg/mL, 77 mg/mL, 88 mg/mL, 99 mg/mL, or 100 mg/mL.
  • the ASO is present in the liquid composition at a concentration of about 22.5 mg/mL, 25 mg/mL, 27.5 mg/mL, 30 mg/mL, 32.5 mg/mL, 35 mg/mL, 37.5 mg/mL, 40 mg/mL, 42.5 mg/mL, 45 mg/mL, 47.5 mg/mL, 50 mg/mL, 52.5 mg/mL, 55 mg/mL, 57.5 mg/mL, 60 mg/mL, 62.5 mg/mL, 65 mg/mL, 67.5 mg/mL, 70 mg/mL, 72.5 mg/mL, 75 mg/mL, 77.5 mg/mL, 80 mg/mL, 82.5 mg/mL, 85 mg/mL, 87.5 mg/mL, 90 mg/mL, 92.5 mg/mL, 95 mg/mL, 97.5 mg/mL, 100 mg/mL, 102.5 mg/mL, 105 mg/mL, 107.5 mg/mL, 110 mg/mL, 52.5 mg/
  • the ASO comprises a sequence with at least 80% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099. [0104] In some embodiments, the ASO comprises a sequence with at least 83%, 88%, 94% or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099. [0105] In some embodiments, the ASO consists of a sequence with at least 83%, 88%, 94% or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the ASO is a compound according to the following chemical structure: WSGR Docket No.47991-744.601 (I), or a salt thereof. [0107] In some embodiments, the ASO is a compound according to the following chemical structure: (II).
  • a liquid pharmaceutical formulation consisting essentially of: (a) an antisense oligomer (ASO) at a concentration of about 0.1 mg/mL to about 500 mg/mL, wherein the ASO is a compound according to the following chemical structure: (I), or a salt thereof; (b) calcium chloride dihydrate (CaCl22H2O) at a concentration of about 0.1 mM to about 50 mM; (c) magnesium chloride hexahydrate (MgCl26H2O) at a concentration of about 0.1 mM to about 50 mM; (d) potassium chloride (KCl) at a concentration of about 0.1 mM to about 20 mM; (e) sodium chloride (NaCl) at a concentration of about 25 mM to about 250 mM; and (f) water.
  • ASO antisense oligomer
  • the pharmaceutical formulation further comprises sodium hydroxide (NaOH) and/or hydrochloric acid (HCl) in amounts that provide a pH of the pharmaceutical formulation at 6.6 to 7.6.
  • the pharmaceutical formulation further comprises sodium hydroxide (NaOH) at a concentration of about 0.1 mM to about 20 mM, and/or hydrochloric acid (HCl) at a concentration of about 0.1 mM to about 20 mM.
  • a liquid pharmaceutical formulation consisting essentially of: (a) an antisense oligomer (ASO) at a concentration of about 0.1 mg/mL to about 500 mg/mL, wherein the ASO is a compound according to the following chemical structure: WSGR Docket No.47991-744.601 (I), or a salt thereof; (b) calcium chloride dihydrate (CaCl22H2O) at a concentration of about 0.1 mM to about 50 mM; (c) magnesium chloride hexahydrate (MgCl26H2O) at a concentration of about 0.1 mM to about 50 mM; (d) potassium chloride (KCl) at a concentration of about 0.1 mM to about 20 mM; (e) sodium chloride (NaCl) at a concentration of about 25 mM to about 250 mM; (f) sodium hydroxide (NaOH) at a concentration of about 0.1 mM to about 20 mM
  • the concentration of the ASO is about 0.1 mg/mL to about 250 mg/mL. In some aspects, the concentration of the ASO is from 6.7 mg/mL to 188 mg/mL, from 6.8 mg/mL to 187 mg/mL, from 3 mg/mL to 100 mg/mL, or from 3 mg/mL to 33 mg/mL.
  • the concentration of the ASO is about 0.1 mg/mL, 0.5 mg/mL, 1 mg/mL, 2 mg/mL, 2.5 mg/mL, 3 mg/mL, 4 mg/mL, 4.5 mg/mL, 5 mg/mL, 6 mg/mL, 7 mg/mL, 8 mg/mL, 9 mg/mL, 10 mg/mL, 11 mg/mL, 12 mg/mL, 13 mg/mL, 14 mg/mL, 15 mg/mL, 16 mg/mL, 17 mg/mL, 18 mg/mL, 19 mg/mL, 20 mg/mL, 22 mg/mL, 25 mg/mL, 28 mg/mL, or 33 mg/mL.
  • the concentration of the ASO is 11 mg/mL, 22 mg/mL, 33 mg/mL, 44 mg/mL, 55 mg/mL, 66 mg/mL, 77 mg/mL, 88 mg/mL, 99 mg/mL, or 100 mg/mL.
  • the concentration of the ASO is about 3 mg/mL, 4 mg/mL, 4.5 mg/mL, 5 mg/mL, 6 mg/mL, 7 mg/mL, 9 mg/mL, 10 mg/mL, 11 mg/mL, 12 mg/mL, 13 mg/mL, 14 mg/mL, 15 mg/mL, 16 mg/mL, 17 mg/mL, 18 mg/mL, 19 mg/mL, 20 mg/mL, 22.5 mg/mL, 25 mg/mL, 27.5 mg/mL, 30 mg/mL, 32.5 mg/mL, 35 mg/mL, 37.5 mg/mL, 40 mg/mL, 42.5 mg/mL, 45 mg/mL, 47.5 mg/mL, 50 mg/mL, 52.5 mg/mL, 55 mg/mL, 57.5 mg/mL, 60 mg/mL, 62.5 mg/mL, 65 mg/mL, 67.5 mg/mL, 70 mg/mL, 72.5 mg/
  • the concentration of calcium chloride dihydrate is 0.2 mM to 25 mM, 0.5 mM to 10 mM, 0.75 mM to 5 mM, or 1 mM to 2 mM. In some aspects, the concentration of calcium chloride dihydrate is from about 1 mM to about 2 mM. In some aspects, the concentration of calcium chloride dihydrate is about 1.4 mM. In some aspects, the concentration of magnesium chloride hexahydrate is 0.2 mM to 25 mM, 0.3 mM to 15 mM, 0.4 mM to 5 mM, 0.5 mM to 1.5 mM, or 0.6 mM to 1 mM.
  • the concentration of magnesium chloride hexahydrate is from about 0.6 mM to about 1 mM. In some aspects, the concentration of magnesium chloride hexahydrate is about 0.79 mM. In some aspects, the concentration of potassium chloride is 0.5 mM to 10 mM, 1 mM to 7.5 mM, or 2 mM to 5 mM. In some aspects, the concentration of potassium chloride is from about 2 mM to about 5 mM. In some aspects, the concentration of potassium chloride is about 3 mM. In some aspects, the concentration of sodium chloride is 25 mM to 250 mM, 100 mM to 160 mM, 110 mM to 140 mM, or 130 mM to 160 mM.
  • the concentration of sodium chloride is from about 125 mM to 145 mM. In some aspects, the concentration of sodium chloride is about 130 mM. In some aspects, the concentration of sodium chloride is from about 140 mM to 160 mM. In some aspects, the concentration of sodium chloride is about 150 mM. In some aspects, the concentration of calcium chloride dihydrate is from about 0.5 mM to about 5 mM; the concentration of magnesium chloride hexahydrate is from about 0.3 mM to about 1.25 mM; the concentration of potassium chloride is from about 1 mM to about 10 mM; and the concentration of sodium chloride is from about 100 mM to 180 mM.
  • the concentration of calcium chloride dihydrate is from about 1 mM to about 2 mM; the concentration of magnesium chloride hexahydrate is from about 0.6 mM to about 1 mM; the concentration of potassium chloride is from about 2 mM to about 5 mM; and the concentration of sodium chloride is from about 120 mM to 160 mM.
  • the concentration of the ASO is about 3 mg/mL, about 4.5 mg/mL, about 7 mg/mL, or about 33 mg/mL; the concentration of calcium chloride dihydrate is about 1.4 mM; the concentration of magnesium chloride hexahydrate is about 0.79 mM; the concentration of potassium chloride is about 3 mM; and the concentration of sodium chloride is about 150 mM.
  • the pH of the pharmaceutical composition is about 6.6 to about 7.6. In some aspects, the pH of the pharmaceutical composition is about 6.8 to about 7.2. In some aspects, the pH of the pharmaceutical composition is about 6.9 to about 7.1.
  • the volume of the pharmaceutical composition is about 5 mL, about 6 mL, about 7 mL, about 8 mL, about 9 mL, about 10 mL, about 12 mL, about 15 mL, WSGR Docket No.47991-744.601 about 20 mL, about 25 mL. In some aspects, the volume of the pharmaceutical composition is about 10 mL.
  • a liquid pharmaceutical formulation consisting essentially of: (a) an antisense oligomer (ASO) at a concentration of about 0.1 mg/mL to about 500 mg/mL, wherein the ASO is a compound according to the following chemical structure: (I), or a salt thereof; (b) calcium ion (Ca 2+ ) at a concentration of about 0.1 mM to about 50 mM; (c) magnesium ion (Mg 2+ ) at a concentration of about 0.1 mM to about 50 mM; (d) potassium ion (K + ) at a concentration of about 0.1 mM to about 20 mM; (e) sodium ion (Na + ) at a concentration of about 25 mM to about 250 mM; (f) chloride ion (Cl-) at a concentration of about 25 mM to about 250 mM; and (g) water.
  • ASO antisense oligomer
  • a liquid pharmaceutical formulation consisting essentially of: (a) an antisense oligomer (ASO) at a concentration of about 0.1 mg/mL to about 500 mg/mL, wherein the ASO is a compound according to the following chemical structure: WSGR Docket No.47991-744.601 (I), or a salt thereof; (b) calcium ion (Ca 2+ ) at a concentration of about 1.4 mM; (c) magnesium ion (Mg 2+ ) at a concentration of about 0.79 mM; (d) potassium ion (K + ) at a concentration of about 3 mM; (e) sodium ion (Na + ) at a concentration of about 160 mM; (f) chloride ion (Cl-) at a concentration of about 160 mM; and (g) water.
  • ASO antisense oligomer
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: i) an active pharmaceutical ingredient (API), and ii) a pharmaceutically acceptable diluent; wherein the liquid composition is not buffered by a buffering agent; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, and/or (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject.
  • API active pharmaceutical ingredient
  • a pharmaceutically acceptable diluent wherein the liquid composition is not buffered by a buffering agent
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion
  • the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising:(i) an active pharmaceutical ingredient (API), and (ii) a pharmaceutically acceptable diluent; wherein the liquid composition lacks Na 2 HPO 4 and/or NaH 2 PO 4 ; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, and/or (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject.
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: (i) an active pharmaceutical ingredient (API), and (ii) a pharmaceutically acceptable diluent consisting of: a) NaCl, b) KCl, c) MgCl 2 or MgCl 2 WSGR Docket No.47991-744.601 6H2O, and d) CaCl2 or CaCl22H2O.
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • the pharmaceutically acceptable diluent provided herein is suitable for preparation of any pharmaceutical formulation intrathecal administration.
  • any active pharmaceutical ingredient suitable for intrathecal administration may be formulated using the pharmaceutically acceptable diluent provided herein.
  • the active pharmaceutical ingredient can be a small molecule or a biologic, such as an organic chemical compound, an antisense oligonucleotide, a DNA, an antibody or any other protein or polypeptide, a living organism such as bacterium or fungi, viral vector, or viral-like particle, or any combination thereof.
  • Also provided herein is a method of treating or reducing the likelihood of developing a disease or condition in a subject in need thereof, the method comprising administering to the subject a pharmaceutical composition disclosed herein.
  • the subject is a human subject.
  • the human subject is at most 18 years old at a first dose of the pharmaceutical composition.
  • the method comprises administering the ASO at a first dose of from about 0.5 milligrams to about 500 milligrams.
  • the method comprises administering multiple doses of the ASO.
  • the disease or condition is characterized by a reduced expression or function of Na V 1.1 protein in the subject.
  • the disease or condition is Dravet Syndrome.
  • the subject is characterized by having: a. seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; b. no past history of causal magnetic resonance imaging lesion; c. no other known etiology of any diseases or conditions except Dravet Syndrome; d. normal development at seizure onset; e. a pathogenic variant, or variant of uncertain significance in an SCN1A gene; f. at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; g.
  • convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures are any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or Atonic (Drop Attacks), and Clonic; h. a current intervention for epilepsy or medication with at least one antiepileptic drug at a dose which has been stable for at least 4 weeks, wherein the interventions for epilepsy is WSGR Docket No.47991-744.601 a ketogenic diet, a vagal nerve stimulator, or a cannabinoid or marijuana-derived product; or i. any combination thereof.
  • the subject is characterized by not having one or more of the following: a. one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; b.
  • a known pathogenic mutation in another gene that causes epilepsy wherein the pathogenic mutation is homozygous in cases of known recessive disease; c. currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide, and wherein the anticoagulant is not an aspirin; d. clinically, significantly unstable medical conditions other than epilepsy; e. clinically, relevant symptoms or a clinically significant illness in the 4 weeks prior to administering, other than epilepsy; f.
  • a history of brain or spinal cord disease other than epilepsy, Dravet Syndrome or a history of bacterial meningitis or brain malformation g. a spinal deformity or other condition that alters the free flow of cerebrospinal fluid (CSF) or having an implanted CSF drainage shunt; h. clinically significant abnormal laboratory values prior to administering; i. aspartate aminotransferase or alanine aminotransferase >2.5-fold upper limit of normal, serum creatinine greater than an upper limit of normal or platelet count less than a lower limit of normal; j. clinically relevant abnormalities in the 12-lead electrocardiogram (ECG) measured at prior to administering; k.
  • ECG electrocardiogram
  • the subject is from 1 to 18, from 2 to 18, from 3 to 18, from 4 to 18, from 5 to 18, from 6 to 18, from 7 to 18, from 8 to 18, from 9 to 18, from 10 to 18, from 11 to 18, from 12 to 18, from 13 to 18, from 14 to 18, from 15 to 18, from 16 to 18, or from 17 to 18 years old.
  • the subject is a human from 1 to 17, from 1 to 16, from 1 to 15, from 1 to 14, from 1 to 13, from 1 to 12, from 1 to 11, from 1 to 10, from 1 to 9, from 1 to 8, from 1 to 7, from 1 to 6, from 1 to 5, from 1 to 4, from 1 to 3, or from 1 to 2 years old.
  • WSGR Docket No.47991-744.601 In some embodiments, the subject is less than a year old or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 years old.
  • the pharmaceutical composition is administered into the intrathecal space of the subject.
  • the pharmaceutical composition is administered into the cerebrospinal fluid of the subject.
  • the pharmaceutical composition is administered into the brain of the subject. [0133] In some embodiments, the pharmaceutical composition is administered into the cerebrospinal fluid in the brain of the subject. [0134] In some embodiments, the pharmaceutical composition is administered as a bolus injection. [0135] In some embodiments, the method comprises administering the pharmaceutical composition as a bolus injection over 1 to 60 minutes, 1 to 50 minutes, 1 to 40 minutes, 1 to 30 minutes, 1 to 20 minutes, 1 to 10 minutes, 1 to 5 minutes, or 1 to 3 minutes. [0136] In some embodiments, the pharmaceutical composition is administered by infusion with a delivery pump. [0137] In some embodiments, the pharmaceutical composition is administered by intracerebroventricular injection.
  • the pharmaceutical composition is administered by intrathecal injection.
  • the method reduces or ameliorates at least one symptom of Dravet Syndrome in the human subject.
  • the symptom of Dravet Syndrome is a seizure.
  • the administration reduces or ameliorates seizure frequency, seizure intensity, or seizure duration.
  • the method further comprises assessing tolerability or effectiveness of the pharmaceutical composition.
  • the method further comprises administering to the subject a pharmaceutical composition comprising the ASO at subsequent doses of from about 0.5 milligrams to about 500 milligrams.
  • the subsequent dose is 0.1, 0.5, 1, 2.5, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 22.5, 25, 27.5, 30, 32.5, 35, 37.5, 40, 42.5, 45, 47.5, 50, 52.5, 55, 57.5, 60, 62.5, 65, 67.5, 70, 72.5, 75, 77.5, 80, 82.5, 85, 87.5, 90, 92.5, 95, 97.5, 100, 102.5, 105, 107.5, 110, 112.5, 115, 117.5, 120, 122.5, 125, 127.5, 130, 132.5, 135, 137.5, 140, 142.5, 145, 147.5, 150, 152.5, 155, 157.5, 160, 162.5, 165, 167.5, 170, 172.5, 175, 177.5, 180, 182.5, 185, 187.5, 190, 192.5, 195, 197.5, 200, 202.5, 205, 207.5, 210, 212.5, 215, 217.5, 220, 222.5, 225,
  • the subsequent dose is lower than the previous dose following an indication that administration of the previous dose is not tolerated.
  • WSGR Docket No.47991-744.601 [0146] In some embodiments, the subsequent dose is the same as the previous dose following an indication that administration of the previous dose is tolerated. [0147] In some embodiments, the subsequent dose is higher than the previous dose following an indication that administration of the previous dose is tolerated. [0148] In some embodiments, the subsequent dose is the same as the previous dose following an indication that administration of the previous dose is effective. [0149] In some embodiments, the subsequent dose is lower than the previous dose following an indication that administration of the previous dose is effective.
  • the subsequent dose is higher than the previous dose following an indication that administration of the previous dose is not effective.
  • the subsequent doses are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months after administration of the previous dose.
  • dose frequency is maintained or reduced following an indication that the previous dose is effective.
  • dose frequency is increased following an indication that the previous dose is not effective.
  • the method further comprises administrating at least one additional therapeutic agent or therapy.
  • the at least one additional therapeutic agent or therapy is administered at the same time as the dose.
  • the at least one additional therapeutic agent or therapy is administered prior to administration of the dose.
  • the at least one additional therapeutic agent or therapy is administered after administration of the dose.
  • the reduced expression or function of Na V 1.1 protein is associated with an altered splicing of a nonsense-mediated RNA decay-inducing exon from a pre-mRNA that contains the NMD exon and encodes Na V 1.1 protein.
  • the ASO promotes exclusion of the NMD exon from the pre-mRNA that contains the NMD exon and that encodes the Na V 1.1 protein.
  • the ASO binds to a targeted portion of a pre-mRNA that contains the NMD exon and that encodes the NaV1.1 protein.
  • the ASO promotes exclusion of the NMD exon from the pre-mRNA that contains the NMD exon and that encodes the NaV1.1 protein.
  • the ASO increases a level of processed mRNA encoding the NaV1.1 protein when the ASO is introduced into the cell.
  • the ASO increases a level of the NaV1.1 protein when the ASO is introduced into the cell.
  • the targeted portion is within an intron sequence flanking the NMD exon.
  • the targeted portion comprises at least one nucleotide of the NMD exon.
  • the targeted portion is within the NMD exon.
  • Also provided herein is a method of making a pharmaceutical composition described herein, the method comprising diluting the antisense oligomer (ASO) in the pharmaceutically acceptable diluent, thereby making the liquid composition.
  • the ASO is present in the liquid composition at a concentration of 3 mg/mL, 4 mg/mL, 4.5 mg/mL, 5 mg/mL, 6 mg/mL, 7 mg/mL, 9 mg/mL, 10 mg/mL, 11 mg/mL, 12 mg/mL, 13 mg/mL, 14 mg/mL, 15 mg/mL, 16 mg/mL, 17 mg/mL, 18 mg/mL, 19 mg/mL, 20 mg/mL, 22 mg/mL, 25 mg/mL, 28 mg/mL, 30 mg/mL, 44 mg/mL, 55 mg/mL, 66 mg/mL, 77 mg/mL, 88 mg/mL, 99 mg/mL, 100 mg/mL, 22.5 mg/mL, 25 mg/mL, 27.5 mg/mL, 30 mg/mL, 32.5 mg/mL, 35 mg/mL, 37.5 mg/mL, 40 mg/mL, 42.5
  • the method further comprises diluting the liquid composition with an additional volume of the pharmaceutically acceptable diluent, thereby making the liquid composition.
  • the ASO is present in the liquid composition at a concentration of wherein the ASO is present in the liquid composition at a concentration of from 0.1-500 mg/mL, from 0.1 mg/mL to 250 mg/mL, from 6.7 mg/mL to 188 mg/mL, from 6.8 mg/mL to 187 mg/mL, from 3 mg/mL to 100 mg/mL, or from 3 mg/mL to 33 mg/mL, or about 0.1 mg/mL, 0.5 mg/mL, 1 mg/mL, 2 mg/mL, 2.5 mg/mL, 3 mg/mL, 4 mg/mL, 4.5 mg/mL, 5 mg/mL, 6 mg/mL, 7 mg/mL, 8 mg/mL, 9 mg/mL, 10 mg/mL, 11 mg/mL
  • the method further comprises filtering the liquid composition.
  • filtering comprises filtering the liquid composition at least twice or through at least two membranes.
  • filtering comprises filtering the liquid composition through a 0.45- ⁇ m membrane and a 0.2- ⁇ m membrane.
  • the at least two membranes, the 0.45- ⁇ m membrane, and/or the 0.2- ⁇ m membrane comprises a polyethersulfone membrane.
  • the method further comprises filling a vial with the liquid composition.
  • the vial is sterilized and/or depyrogenated.
  • the method further comprises fitting the vial with a stopper. [0178] In some embodiments, the stopper is sterilized and/or depyrogenated. [0179] In some embodiments, the method further comprises capping the vial with a seal. [0180] In some embodiments, the seal is sterilized and/or depyrogenated. [0181] In some embodiments, the method if performed aseptically. [0182] In some embodiments, the method further comprises storing the vial for a time period at a temperature. [0183] In some embodiments, the temperature is -20 ⁇ 5 °C. [0184] In some embodiments, the time period is no more than 36 months.
  • the time period is no more than 24 months.
  • the method further comprises administering the liquid composition to a human subject after the storing.
  • FIGs.1A-1B depict a schematic representation of a target pre-mRNA that contains a nonsense- mediated RNA decay-inducing exon (NMD exon mRNA) and therapeutic agent-mediated exclusion of the nonsense-mediated mRNA decay-inducing exon from the pre-mRNA to increase expression of the full-length target protein or functional RNA.
  • FIG.1A shows a cell divided into nuclear and cytoplasmic compartments.
  • a pre-mRNA transcript of a target gene undergoes splicing to generate processed mRNA, and this processed mRNA is exported to the cytoplasm and translated into target protein.
  • some fraction of the processed mRNA contains a nonsense-mediated mRNA decay-inducing exon (NMD exon mRNA) that is degraded in the cytoplasm, thus leading to no target protein production.
  • NMD exon mRNA nonsense-mediated mRNA decay-inducing exon
  • FIG.1B shows an example of the same cell divided into nuclear and cytoplasmic compartments.
  • FIG.1C is a schematic representation of therapeutic ASO-mediated exclusion of a nonsense- mediated mRNA decay-inducing exon from a pre-mRNA, which decreases non-productive processed WSGR Docket No.47991-744.601 mRNA (e.g., with an NMD exon) and increases productive mRNA (e.g., without an NMD exon) and increases expression of the full-length target protein from the productive mRNA.
  • a therapeutic agent such as an antisense oligomer (ASO)
  • FIG.1D shows identification of an exemplary sequence in the SCN1A gene that encodes a nonsense-mediated mRNA decay (NMD)-inducing exon.
  • the identification of the sequence in the SCN1A gene that encodes the NMD-inducing exon using comparative genomics is shown, visualized in the UCSC genome browser.
  • the upper panel shows a graphic representation of the SCN1A gene to scale. The conservation level across 100 vertebrate species is shown as peaks. The highest peaks correspond to exons (black boxes), while no peaks are observed for the majority of the introns (lines with arrow heads). Peaks of conservation were identified in intron 20 (NM_006920), shown in the middle panel.
  • FIG.2 shows a study design timeline for monitoring wild type (WT) and Dravet Syndrome (DS) mice as well as a Kaplan-Meier curve showing DS and WT littermate mice monitored to 14 weeks for survival.
  • FIG.3 shows an experimental design for the EEG seizure monitoring study in DS mice and their WT littermates.
  • FIGs.4A-4E show the results of monitoring seizures in mice administered with ASO-22 or phosphate buffered saline (PBS).
  • FIG.4A shows exemplary EEG recordings in DS mice.
  • FIG.4B shows the number of seizures occurring in various regions of the brain in the two mice groups. *Indicates p ⁇ 0.05.
  • FIG.4D shows the number of mice that had a number of seizures in each group.
  • FIGs.5A-5G show that a single ICV injection of 20 ⁇ g ASO-22 at P2 results in reduced sudden unexpected death in epilepsy (SUDEP) incidence and increased Na V 1.1 protein expression in DS mice.
  • FIG.5A is a schematic for the experimental design.
  • FIGs.5B, 5C, 5D, 5E, 5F, and 5G illustrate the concentration of ASO-22, fold change in Scn1a gene expression, and NaV1.1 expression in brain tissues at 7 weeks or 14 weeks after a single ICV injection of ASO-22 (20 ⁇ g) or PBS at P2, respectively.
  • FIGs.6A-6B show the percent survival of DS and WT mice after a single ICV injection of ASO- 22 (60 ⁇ g) or PBS at P14.
  • FIGs.7A-7F show the ASO-22 exposure, Scn1a expression, and NaV1.1 expression in brain tissues at P35 and P90 after a single ICV injection of ASO-22 (60 ⁇ g) or PBS on P14, respectively.
  • FIG.8 shows the experimental conditions and numbers of monkeys used per group.
  • FIGs.9A-9B show the levels of ASO-22 in various regions of cynomolgus monkey brain on study day 3 and day 29, respectively.
  • WSGR Docket No.47991-744.601 [0200]
  • FIGs.10A-10B show the levels of NaV1.1 protein in various regions of cynomolgus monkey brain on study day 3 and day 29, respectively.
  • FIGs.11A-11B show the percentages of productive SCN1A gene to total SCN1A gene as an evaluation of target engagement in cynomolgus monkeys on day 3 and day 29.
  • FIG.12A shows the plasma pharmacokinetics in cynomolgus monkey at various timepoints after intrathecal administration of ASO-22.
  • FIG.12B shows the levels of ASO-22 in the cynomolgus monkey cerebrospinal fluid (CSF) on study day 3 and 29.
  • FIGs.13A-13D depict identification of an alternative splicing event in SCN1A that results in NMD.
  • FIG.13A shows SCN1A splicing isoforms with or without inclusion of the alternative exon in ReNcells as demonstrated by RT-PCR.
  • FIG.13B shows evaluation of the alternative splice event of the SCN1A gene in the cerebral cortex of four species.
  • FIG.13C shows an image of a TBE PAGE gel of RT- PCR products corresponding to Scn1a productive (lower bands, 498 bp) and non-productive transcript (upper bands, 562 bp) amplified from total RNA extracted from WT C57BL/6J mouse brains from P0 to P20 and at 10 months.
  • Mouse Gapdh was used as a loading control.
  • FIG.13D summarizes expression of Scn1a productive and non-productive transcript in postnatal mouse brains, calculated with optical densities of PCR products shown in FIG.13C.
  • FIGs.14A-14E illustrate that selected ASOs suppressed the NMD splicing event and increased the expression of productive SCN1A mRNA in ReNcells.
  • FIGs.15A-15C show dose-dependent effects of ASO-22 on splicing and expression of SCN1A mRNA in ReNcells.
  • FIGs.16A-16H illustrate that ASO-22 ICV injection causes dose-dependent and durable increases in productive Scn1a mRNA and Na V 1.1 protein expression in mouse brain.
  • FIG.18 shows dose-dependent effects of ASO-22 on the expression of Na V 1.1 in ICV-injected neonatal mouse brains.
  • FIG.20 shows expression of Na V 1.1 in mouse brains at different post-injection days.
  • FIG.21 shows validation of the two anti-NaV1.1 antibodies used in the Examples. Specificity of the two anti-NaV1.1 antibodies, Alomone ASC-001 and NeuroMab 75-023, was tested using total protein prepared from a Scn1a -/- mouse brain (middle lane) and brains of two WT littermates (left and right lanes).
  • FIG.22 shows a schematic representation of clinical manifestations of Dravet Syndrome and their relative incidences according to age.
  • AA atypical absences
  • AE acute encephalopathy
  • CG crouching gait
  • CPS complex partial seizures
  • DD developmental delay
  • DS Dravet syndrome
  • EEG electroencephalogram
  • FSz complex febrile seizures
  • GMS generalized motor seizures
  • HS hyperthermia sensitivity
  • m month
  • MSz myoclonic seizures
  • OS obtundation status
  • SE status WSGR Docket No.47991-744.601 epilepticus
  • SUDEP sudden unexpected death in epilepsy
  • y years
  • FIG.23 shows TANGO (Targeted Augmentation of Nuclear Gene Output) that may be used to treat Dravet syndrome.
  • FIG.24 shows the transformative potential of TANGO technology in Dravet syndrome.
  • FIG.25 shows the study design. Phase 1/2a open-label, two-part study conducted at approximately 20 sites in the United States.
  • FIG.26 shows a schematic representation of the study design.
  • FIG.27 shows patient inclusion and exclusion criteria.
  • FIG.28 shows study assessments.
  • FIG.29 shows the effect of phosphate on the pH of the ASO-1 formulation.
  • FIG.30 shows oligonucleotide self-buffering capacity vs pH for the titrations of ASO-1 at 33 mg/mL and 5 mg/mL.
  • FIG.31 shows the readout of Fourier-transform infrared spectroscopy (FTIR).
  • FIG.32 shows the Scanning electron microscopy-energy dispersive X-ray spectroscopy (SEM- EDS).
  • FIG.33 is a representative photograph of fiber-like particles present in an ASO formulation at 1 and 3 months when stored at 25 °C.
  • This exemplary formulation contains 4.58 mM ASO-1, 1.4 mM CaCl 2 , 0.79 mM MgCl 2 , 3 mM KCl, 150 mM NaCl, 0.70 mM Na 2 HPO 4 ⁇ 2H 2 O, and 0.3 mM NaH2PO4, ⁇ 2H2O at pH 7.0 to 7.5.
  • FIG.34 is a representative photograph of large, waxy-looking flakes that were observed in stability tests of formulations containing either 4.58 mM ASO-1, 1.4 mM CaCl 2 , 0.79 mM MgCl 2 , 3 mM KCl, 0.70 mM Na2HPO4 ⁇ 2H2O, and 0.3 mM NaH2PO4, ⁇ 2H2O; or containing 4.58 mM ASO-1, 1.4 mM CaCl 2 , 0.79 mM MgCl 2 , 3 mM KCl, 150 mM NaCl, 0.70 mM Na 2 HPO 4 ⁇ 2H 2 O, and 0.3 mM NaH2PO4, ⁇ 2H2O but without pH adjustment.
  • the following phrases “A, B, and/or C” or “A, B, C, or any combination thereof” can mean “A individually; B individually; C individually; A and B; B and C; A and C; and A, B, and C.”
  • the term “or” can be used conjunctively or disjunctively, unless the context specifically refers to a disjunctive use.
  • the term “about” or “approximately” can mean within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, i.e., the limitations of the measurement system.
  • “about” can mean within 1 or more than 1 standard deviation, per the practice in the art. Alternatively, “about” can mean a range of up to 20%, up to 10%, up to 5%, or up to 1% of a given value. Alternatively, particularly with respect to biological systems or processes, the term can mean within an order of magnitude, within 5-fold, and more preferably within 2-fold, of a value. Where particular values are described in the application and claims, unless otherwise stated, the term “about” meaning within an acceptable error range for the particular value should be assumed.
  • the words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or “containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open-ended and do not exclude additional, unrecited elements or method steps. It is contemplated that any embodiment discussed in this specification can be implemented with respect to any method or composition of the present disclosure, and vice versa. Furthermore, compositions of the present disclosure can be used to achieve methods of the present disclosure.
  • a nucleic acid sequence is a polymer comprising or consisting of nucleotide monomers, which are covalently linked to WSGR Docket No.47991-744.601 each other by phosphodiester-bonds of a sugar/phosphate-backbone.
  • the term “nucleic acid sequence” also encompasses modified nucleic acid sequences, such as base-modified, sugar-modified or backbone- modified, etc., DNA or RNA.
  • fragment or “fragment of a sequence” which have the identical meaning herein is a shorter portion of a full-length sequence of e.g., a nucleic acid molecule like DNA or RNA or a protein.
  • a fragment typically, consists of a sequence that is identical to the corresponding stretch within the full-length sequence.
  • a preferred fragment of a sequence in the context of the present invention consists of a continuous stretch of entities, such as nucleotides or amino acids corresponding to a continuous stretch of entities in the molecule the fragment is derived from, which represents at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, at least 7%, at least 8%, at least 9%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, 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%, at least 97%, at least 98%, at least 99%, at least 99.5%, or even 100% of the total (i.e., full-length) molecule from which the fragment is
  • a “fragment” or “functional fragment” of a polynucleotide or a polypeptide is a fragment of the polynucleotide or the polypeptide that is shorter than the full-length, immature, or mature nucleotide or polypeptide and has at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, at least 7%, at least 8%, at least 9%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, 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%, at least 97%, at least 98%, at least 99%, at least 99.5%, or even 100% or more of the activity of the full-length mature reference polynucleotide or polypeptide.
  • Fragments of interest can be made by recombinant, synthetic, or digestive methods.
  • the term “recombinant polynucleotide” can refer to a polynucleotide that is not naturally occurring and are synthesized or manipulated in vitro, such as polynucleotides produced by laboratory methods.
  • a recombinant polynucleotide can be synthesized in a laboratory and/or can be prepared by using recombinant DNA technology by using enzymatic modification of DNA, such as enzymatic restriction digestion, ligation, and cloning.
  • a recombinant polypeptide can be prepared by in vitro transcription of a recombinant DNA followed by in vitro translation of the produced messenger RNA (mRNA).
  • mRNA messenger RNA
  • a recombinant polynucleic acid or RNA can be incorporated into a cell and a recombinant polypeptide can be expressed within the cell.
  • Recombinant proteins can include amino acid residues not found within the native (non-recombinant) form of the protein or can include amino acid residues that have been modified, e.g., labeled.
  • isolated means separated from constituents, cellular and otherwise, in which the polynucleotide, polypeptide, protein, or fragments thereof, are normally associated with in nature.
  • an isolated polynucleotide is one that is separated from the 5’ WSGR Docket No.47991-744.601 and 3’ ends with which it is normally associated in the naturally occurring sequence.
  • nucleotide means a nucleoside further comprising a phosphate linking group.
  • nucleosides may or may not be linked by phosphate linkages and thus includes, but is not limited to, “linked nucleotides.”
  • linked nucleosides are nucleosides that are connected in a continuous sequence (i.e., no additional nucleosides are present between those that are linked).
  • nucleobase means a group of atoms that can be linked to a sugar moiety to create a nucleoside that is capable of incorporation into an oligonucleotide, and wherein the group of atoms is capable of bonding with a complementary naturally occurring nucleobase of another oligonucleotide or nucleic acid.
  • Nucleobases may be naturally occurring or may be modified.
  • nucleoside means a compound comprising a nucleobase moiety and a sugar moiety. Nucleosides include, but are not limited to, naturally occurring nucleosides (as found in DNA and RNA) and modified nucleosides. Nucleosides may be linked to a phosphate moiety.
  • naturally occurring sugar moiety means a ribofuranosyl as found in naturally occurring RNA or a deoxyribofuranosyl as found in naturally occurring DNA.
  • sugar moiety means a naturally occurring sugar moiety or a modified sugar moiety of a nucleoside.
  • modified sugar moiety means a substituted sugar moiety, a bicyclic or tricyclic sugar moiety, or a sugar surrogate.
  • antisense oligonucleotide refers to synthetic antinucleotide oligonucleotide (ASO) or antisense oligonucleotide analogs usually between 12 and 30 nucleotides in length that are designed to hybridize to RNA by Watson-Crick base pairing.
  • ASOs can be designed to bind to protein coding RNAs (mRNAs) as well as noncoding RNAs such as microRNAs or large noncoding RNAs. After binding to the targeted RNA, the ASO can modulate the function of the targeted RNA by several different mechanisms, including degradation of the pre-mRNA in the nucleus or mature RNA in the cytoplasm by RNase H1, and degradation of RNA in the cytoplasm by the RISC complex (Ago2) or ribozymes or DNAzymes. ASOs can also modulate RNA function by nondegradative mechanisms such as splicing or polyadenylation modulation in the nucleus and modulate protein translation in the cytoplasm.
  • mRNAs protein coding RNAs
  • noncoding RNAs such as microRNAs or large noncoding RNAs.
  • to hybridize means to form hydrogen bond, which may be via Watson-Crick, Hoogsteen or reversed Hoogsteen hydrogen bonding, between complementary nucleoside or nucleotide bases.
  • “Complementary,” as used herein, refers to the capacity for precise pairing between two WSGR Docket No.47991-744.601 nucleotides. The oligonucleotide and the DNA or RNA are complementary to each other when a sufficient number of corresponding positions in each molecule are occupied by nucleotides which can hydrogen bond with each other.
  • nucleic acids or polypeptide sequences refer to two or more sequences or subsequences that are the same or have a specified percentage of nucleotides or amino acid residues that are the same (i.e., 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100%identity over a specified region, e.g., of the entire polypeptide sequences of the invention or individual domains of the polypeptides of the invention), when compared and aligned for maximum correspondence over a comparison window, or designated region as measured using a sequence comparison algorithm or by manual alignment and visual inspection.
  • sequences that are at least about 80% identical are said to be “substantially identical.”
  • two sequences are 100% identical.
  • two sequences are 100% identical over the entire length of one of the sequences (e.g., the shorter of the two sequences where the sequences have different lengths).
  • identity may refer to the complement of a test sequence. [0245] In some embodiments, the identity exists over a region that is at least about 2 to about 400 amino acids or nucleotides in length.
  • the identity exists over a region that is at least about 2 to about 390, at least about 2 to about 380, at least about 2 to about 370, at least about 2 to about 360, at least about 2 to about 350, at least about 2 to about 340, at least about 2 to about 330, at least about 2 to about 320, at least about 2 to about 310, at least about 2 to about 300, at least about 2 to about 290, at least about 2 to about 280, at least about 2 to about 270, at least about 2 to about 260, at least about 2 to about 250, at least about 2 to about 200, at least about 2 to about 150, at least about 2 to about 100 amino acids or nucleotides in length.
  • the identity exists over a region that is at least about 2 to about 90, at least about 2 to about 85, at least about 2 to about 80, at least about 2 to about 75, at least about 2 to about 70, at least about 2 to about 65, at least about 2 to about 60, at least about 2 to about 55, at least about 2 to about 50, at least about 2 to about 45, at least about 2 to about 40, at least about 2 to about 35, at least about 2 to about 30, at least about 2 to about 25, at least about 2 to about 20, at least about 2 to about 10, at least about 2 to about 5 amino acids or nucleotides in length.
  • the identity exists over a region that is at least about 3 to about 400, about 4 to about 400, about 5 to about 400, about 6 to about 400, about 7 to about 400, about 8 to about 400, about 9 to about 400, about 10 to about 400, about 11 to about 400, about 12 to about 400, about 13 to about 400, about 14 to about 400, about 15 to about 400, about 16 to about 400, about 17 to about 400, about 18 to about 400, about 19 to about 400, about 20 to about 400, about 21 to about 400, about 22 to about 400, about 23 to about 400, about 24 to about 400, about 25 to about 400, about 26 to about 400, about 27 to about 400, about 28 to about 400, about 29 to about 400, about 30 to about 400, about 31 to about 400, about 32 to about 400, about 33 to about 400, about 34 to about 400, about 35 to about 400 amino acids or nucleotides in length.
  • the identity exists over a region that is at least about 40 to about 400, about 45 to about 400, about 50 to about 400, about 55 to about 400, about WSGR Docket No.47991-744.601 60 to about 400, about 61 to about 400, about 62 to about 400, about 63 to about 400, about 64 to about 400, about 65 to about 400, about 66 to about 400, about 67 to about 400, about 68 to about 400, about 69 to about 400, about 70, to about 400, about 71 to about 400, about 72 to about 400, about 73 to about 400, about 74 to about 400, about 75 to about 400, about 80 to about 400, about 85 to about 400, about 90 to about 400, about 100 to about 400, about 150 to about 400, about 200 to about 400, about 250 to about 400, about 300 to about 400, about 350 to about 400 amino acids or nucleotides in length.
  • the identity exists over a region that is at least about 2 to about 343, about 3 to about 343, about 4 to about 343, about 7 to about 343, about 9 to about 343, about 11 to about 343, about 15 to about 343, about 16 to about 343, about 20 to about 343, about 25 to about 343, about 62 to about 343, about 2 to about 317, about 3 to about 317, about 4 to about 317, about 7 to about 317, about 9 to about 317, about 11 to about 317, about 15 to about 317, about 16 to about 317, about 20 to about 317, about 25 to about 317, about 62 to about 317, about 2 to about 300, about 3 to about 300, about 4 to about 300, about 7 to about 300, about 9 to about 300, about 11 to about 300, about 15 to about 300, about 16 to about 300, about 20 to about 300, about 25 to about 300, about 62 to about 300, about 2 to about 62, about 3 to about 62, about 4 to about 62, about 7
  • the term “genetically modified” means containing and/or expressing a foreign gene or nucleic acid sequence which in turn, modifies the genotype or phenotype of the cell or its progeny. In other words, it refers to any addition, deletion, or disruption to a cell's endogenous nucleotides.
  • the term “operably linked” can refer to a functional relationship between two or more nucleic acid sequences, e.g., a functional relationship of a transcriptional regulatory or signal sequence to a transcribed sequence.
  • a target motif or a nucleic acid encoding a target motif is operably linked to a coding sequence if it is expressed as a preprotein that participates in targeting the polypeptide encoded by the coding sequence to a cell membrane, intracellular, or an extracellular compartment.
  • a signal peptide or a nucleic acid encoding a signal peptide is operably linked to a coding sequence if it is expressed as a preprotein that participates in the secretion of the polypeptide encoded by the coding sequence.
  • a promoter is operably linked if it stimulates or modulates the transcription of the coding sequence.
  • mammals include, but are not limited to, any member of the mammalian class: humans, non-human primates such as chimpanzees, and other apes and monkey species; farm animals such as cattle, horses, sheep, goats, swine; domestic animals such as rabbits, dogs, and cats; laboratory animals including rodents, such as rats, mice and guinea pigs, and the like.
  • the mammal is a human.
  • the term “animal” as used herein comprises human beings and non-human animals.
  • a “non-human animal” is a mammal, for example, a rodent such as rat or a mouse.
  • a non-human animal is a mouse.
  • compositions are a pharmaceutical formulation, wherein the pharmaceutical formulation is a liquid composition comprising: (i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent; wherein the liquid composition is not buffered by a buffering agent; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • ASO antisense oligomer
  • a pharmaceutically acceptable diluent wherein the liquid composition is not buffered by a buffering agent; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid,
  • the pharmaceutical formulation is a liquid composition comprising: (i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent, wherein the ASO is stable in the pharmaceutical formulation for at least 1 or 2 years at 4 °C.
  • ASO antisense oligomer
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion
  • the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: (i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent; wherein the liquid composition lacks Na 2 HPO 4 and/or NaH 2 PO 4 ; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: (i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent consisting of: a) NaCl, b) KCl, c) MgCl2 or MgCl26H2O, and d) CaCl2 or CaCl22H2O.
  • the liquid composition comprises 0.1-50 mM CaCl2 or CaCl22H2O.
  • the liquid composition comprises 1-2 mM CaCl2 or CaCl22H2O. [0261] In some embodiments, the liquid composition comprises about 1.4 mM CaCl2 or CaCl22H2O. [0262] In some embodiments, the liquid composition comprises 0.1-50 mM MgCl2 or MgCl26H2O. [0263] In some embodiments, the liquid composition comprises 0.5-1.5 mM MgCl2 or MgCl26H2O. [0264] In some embodiments, the liquid composition comprises about 0.79 mM MgCl2 or MgCl26H2O.
  • the liquid composition comprises 5-250 mM NaCl, 0.1-20 mM KCl, 0.1- 50 mM CaCl2 or CaCl22H2O, and 0.1-50 mM MgCl2 or MgCl26H2O.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: (i) an antisense oligomer (ASO), and (ii) a pharmaceutically acceptable diluent; wherein the liquid composition lacks calcium ions and/or magnesium ions; and wherein: (a) the liquid composition comprises potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • ASO antisense oligomer
  • a pharmaceutically acceptable diluent wherein the liquid composition lacks calcium ions and/or magnesium ions
  • the liquid composition comprises potassium ion
  • the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • the liquid composition is formulated for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject.
  • the liquid composition is formulated for administration into cerebrospinal fluid in a brain of a human subject.
  • the ASO comprises at least one modified sugar moiety.
  • each nucleotide of the antisense oligomer comprises a modified sugar moiety.
  • kits comprising: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; and wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO, and wherein the liquid composition is not buffered by a buffering agent; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • ASO antisense oligomer
  • a pharmaceutically acceptable diluent wherein the concentrate is miscible with the pharmaceutically acceptable diluent
  • mixing the ASO with the pharmaceutically acceptable diluent results in
  • kits comprising: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; and wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO, and wherein the ASO is stable in the liquid composition for at least 1 or 2 years at 4 °C.
  • ASO antisense oligomer
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion
  • the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • a kit comprising: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; and wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO, wherein the liquid composition lacks Na2HPO4 and/or NaH2PO4; and wherein: (a) the liquid composition comprises calcium ion, magnesium WSGR Docket No.47991-744.601 ion, and/or potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • ASO antisense oligomer
  • a pharmaceutically acceptable diluent wherein the concentrate is miscible with the pharmaceutically acceptable dil
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • a kit comprising: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent consisting of: a) NaCl, b) KCl, c) MgCl2 or MgCl2 6H2O, and d) CaCl2 or CaCl22H2O, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; and wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO.
  • ASO antisense oligomer
  • a kit comprising: (i) a concentrate comprising an antisense oligomer (ASO); and (ii) a pharmaceutically acceptable diluent, wherein the concentrate is miscible with the pharmaceutically acceptable diluent; wherein mixing the ASO with the pharmaceutically acceptable diluent results in a liquid composition comprising the ASO, wherein the liquid composition lacks calcium ions and/or magnesium ions; and wherein: (a) the liquid composition comprises potassium ion, (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject, and/or (c) each nucleobase of the ASO comprises a modified sugar moiety.
  • ASO antisense oligomer
  • the liquid composition is formulated for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject. [0280] In some embodiments, the liquid composition is formulated for administration into cerebrospinal fluid in a brain of a human subject. [0281] In some embodiments, the ASO comprises at least one modified sugar moiety. [0282] In some embodiments, each nucleotide of the antisense oligomer comprises a modified sugar moiety. [0283] In some embodiments, the kit further comprises (iii) instructions for diluting or solubilizing the ASO in the pharmaceutically acceptable diluent.
  • the liquid composition comprises 25-250 mM NaCl. [0285] In some embodiments, the liquid composition comprises 0.1-20 mM KCl. [0286] In some embodiments, the liquid composition comprises 2-4 mM KCl. [0287] In some embodiments, the liquid composition about 3 mM KCl. [0288] In some embodiments, the liquid composition comprises 100-160 mM NaCl. [0289] In some embodiments, the liquid composition comprises 125-145 mM NaCl. [0290] In some embodiments, the liquid composition comprises about 130 mM NaCl. [0291] In some embodiments, the liquid composition comprises a buffered (pH 6.6 – 7.6) solution.
  • the liquid composition comprises 0.1-50 mM Na2HPO4.
  • the liquid composition comprises 0.1-50 mM NaH2PO4.
  • WSGR Docket No.47991-744.601 [0294] In some embodiments, the liquid composition comprises 5-250 mM NaCl, 0.1-20 mM KCl, 0.1- 50 mM Na2HPO4, and 0.1-50 mM NaH2PO4.
  • the ASO is solubilized in the pharmaceutically acceptable diluent.
  • the pharmaceutically acceptable diluent is an isotonic solution.
  • the ASO is not substantially polydisperse.
  • the liquid composition has an osmolality of less than 150 mM.
  • the liquid composition has an osmolality of about 130 mM.
  • the liquid composition further comprises a carbohydrate.
  • the carbohydrate comprises D-glucose.
  • the liquid composition further comprises 1-100 mM D-glucose.
  • the liquid composition further comprises an antioxidant.
  • the antioxidant is t-butylhydroxyquinoline (TBHQ), butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), vitamin E, or any combination thereof.
  • TBHQ t-butylhydroxyquinoline
  • BHT butylated hydroxytoluene
  • BHA butylated hydroxyanisole
  • vitamin E vitamin E
  • the ASO as described herein is solubilized or diluted in an artificial cerebral spinal fluid (aCSF) solution.
  • aCSF artificial cerebral spinal fluid
  • aCSF for instance, closely matches the electrolyte concentrations and physiological compatibility of endogenous CSF to enable a vital environment for neuronal tissue by maintaining the homeostasis, osmolarity, and pH at physiological levels.
  • isotonic solution refers to a solution that contains an electrolyte balance similar to plasma in the bloodstream. Administration of an isotonic solution to a subject or patient may increase the fluid volume of the subject or patient without a fluid shift.
  • Exemplary isotonic solutions include, but are not limited to 0.9% normal saline, lactated Ringer’s solution, Ringer’s solution, plasma-lyte, and 5% Dextrose in water (D 5 W).
  • hypotonic solution refers to a solution that has a lower concentration of electrolytes than plasma.
  • Administration of a hypotonic solution may lead to shifting fluid out of the bloodstream to the area of higher concentration in the interstitial and intracellular spaces.
  • exemplary hypotonic solutions include, but are not limited to, 0.45% normal saline (half normal saline), 0.33% NaCl solution, 0.225% NaCl solution, and 2.5% Dextrose in water (D2.5W).
  • hypotonic solution refers to a solution that has a higher concentration of electrolytes than plasma.
  • hypertonic solutions may shift fluid from the interstitial and intracellular spaces into the bloodstream to dilute the electrolytes.
  • exemplary hypertonic solutions include, but are not limited to, 3% NaCl solution, 5% Dextrose in 0.45% NaCl (D5 1 ⁇ 2 NS), 5% Dextrose in 0.9% normal saline (D5NS), 5% Dextrose in WSGR Docket No.47991-744.601 lactated Ringer’s solution (D5LR), 10% Dextrose in water (D10W), 20% Dextrose in water (D20W), and 50% Dextrose in water (D50W).
  • the ASO as described herein is solubilized or diluted in a phosphate- buffered (pH 6.6-7.6) solution. In some embodiments, the ASO as described herein is solubilized or diluted in a phosphate-buffered (pH 6.0-8.0) solution. In some embodiments, the ASO as described herein is solubilized or diluted in a phosphate-buffered (pH 5.0-8.0) solution.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 4.5-8.5, pH 4.6-8.5, pH 4.7-8.5, pH 4.8-8.5, pH 4.9-8.5, pH5.0-8.5, pH 5.1-8.5, pH5.2-8.5, pH 5.3-8.5, pH5.4-8.5, pH 5.5-8.5, pH5.6-8.5, pH 5.7-8.5, pH 5.8-8.5, H 5.9-8.5, pH 6.0-8.5, pH 6.1-8.5, pH 6.2-8.5, pH 6.3-8.5, pH 6.4-8.5, pH 6.5-8.5, pH 6.6-8.5, pH 6.7-8.5, pH 6.8-8.5, pH 6.9-8.5, pH 7.0-8.5, pH 7.1-8.5, pH 7.2-8.5, pH 7.3-8.5, pH 7.4-8.5, pH 7.5-8.5, pH 7.6-8.5, pH 7.7-8.5, pH 7.8-8.5, pH 7.9-8.5, pH 8.0-8.5, pH 8.1- 8.5, pH 8.2-8.5,
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 4.5-8.3, pH 4.5-8.2, pH 4.5-8.1, pH 4.5- 8.0, pH 4.5-7.9, pH 4.5-7.8, pH 4.5-7.7, pH 4.5-7.6, pH 4.5-7.5, pH 4.5-7.4, pH 4.5-7.3, pH 4.5-7.2, pH 4.5-7.1, pH 4.5-7.0, pH 4.5-6.9, pH 4.5-6.8, pH 4.5-6.7, pH 4.5-6.6, pH 4.5-6.5, pH 4.5-6.4, pH 4.5-6.3, pH 4.5-6.2, pH 4.5-6.1, pH 4.5-6.0, pH 4.5-5.9, pH 4.5-5.8, pH 4.5-5.7, pH 4.5-5.6, pH 4.5-5.5, pH 4.5- 5.4, pH 4.5-5.3, pH 4.5-5.2, pH 4.5-5.1, pH 4.5-5.0, pH 4.5-4.9, pH 4.5-4.8, pH 4.5-4.7, or pH
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-7.6, pH 6.1-7.6, pH 6.2-7.6, pH 6.3-7.6, pH 6.4-7.6, pH 6.5-7.6, pH 6.6-7.6, pH 6.7- 7.6, pH 6.8-7.6, pH 6.9-7.6, pH 7.0-7.6, pH 7.1-7.6, pH 7.2-7.6, pH 7.3-7.6, pH 7.4-7.6, or pH 7.5-7.6.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.6-8.0, pH 6.6-7.9, pH 6.6-7.8, pH 6.6-7.7, pH 6.6-7.6, pH 6.6-7.5, pH 6.6-7.4, pH 6.6- 7.3, pH 6.6-7.2, pH 6.6-7.1, pH 6.6-7.0, pH 6.6-6.9, pH 6.6-6.8, or pH 6.6-6.7.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-8.0, pH 6.1-8.0, pH 6.2-8.0, pH 6.3-8.0, pH 6.4-8.0, pH 6.5-8.0, pH 6.6-8.0, pH 6.7-8.0, pH 6.8-8.0, pH 6.9-8.0, pH 7.0-8.0, pH 7.1-8.0, pH 7.2-8.0, pH 7.3-8.0, pH 7.4-8.0, pH 7.5-8.0, pH 7.6-8.0, pH 7.7-8.0, pH 7.8- 8.0, or pH 7.9-8.0.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-7.9, pH 6.0-7.8, pH 6.0-7.7, pH 6.0-7.6, pH 6.0-7.5, pH 6.0-7.4, pH 6.0-7.3, pH 6.0-7.2, pH 6.0-7.1, pH 6.0-7.0, pH 6.0-6.9, pH 6.0-6.8, pH 6.0-6.7, pH 6.0-6.6, pH 6.0- 6.5, pH 6.0-6.4, pH 6.0-6.3, pH 6.0-6.2, or pH 6.0-6.1.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 5.7-8.5, 5.8-8.4, 5.9-8.3, 6.0-8.2, 6.1-8.1, 6.2-8.0, 6.3-7.9, 6.4-7.8, 6.5-7.7, or 6.6-7.6.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH about 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0.6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, or 8.0.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0.6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, or 8.0.
  • WSGR Docket No.47991-744.601 [0311]
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-250 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-250, 30-250, 35-250, 40-250, 45-250, 50-250, 55-250, 60-250, 65-250, 70-250, 75-250, 80-250, 85-250, 90-250, 95-250, 100-250, 105-250, 110-250, 115-250, 120-250, 125-250, 130-250, 135- 250, 140-250, 145-250, 150-250, 155-250, 160-250, 165-250, 170-250, 175-250, 180-250, 185-250, 190- 250, 195-250, 200-250, 205-250, 210-250, 215-250, 220-250, 225-250, 230-250, 235-250, 240-250, or 245-250 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-245, 25-240, 25-235, 25-230, 25-225, 25-220, 25-215, 25-210, 25-205, 25-200, 25- 195, 25-190, 25-185, 25-180, 25-175, 25-170, 25-165, 25-160, 25-155, 25-150, 25-145, 25-140, 25-135, 25-130, 25-125, 25-120, 25-115, 25-110, 25-105, 25-110, 25-105, 25-100, 25-95, 25-90, 25-85, 25-80, 25-75, 25-70, 25-65, 25-60, 25-55, 25-50, 25-45, 25-40, 25-35, or 25-30 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 30-245, 35- 240, 40-235, 45-230, 50-225, 55-220, 60-215, 65-210, 70-205, 75-200, 80-195, 85-190, 90-185, 95-180, 100-175, 105-170, 110-165, 115-160, 120-155, 125-150, 130-145 or 135-140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 100-140, 101- 140, 102-140, 103-140, 104-140, 105-140, 106-140, 107-140, 108-140, 109-140, 110-140, 111-140, 112- 140, 113-140, 114-140, 115-140, 116-140, 117-140, 118-140, 119-140, 120-140, 121-140, 122-140, 123- 140, 124-140, 125-140, 126-140, 127-140, 128-140, 129-140, 130-140, 131-140, 132-140, 133-140, 134- 140, 135-140, 136-140, 137-140, 138-140, or 139-140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 100-139, 100-138, 100-137, 100-136, 100-135, 100-134, 100-133, 100-132, 100-131, 100-130, 100-129, 100-128, 100-127, 100-126, 100-125, 100-124, 100-123, 100-122, 100-121, 100-120, 100-119, 100-118, 100-117, 100-116, 100-115, 100-114, 100-113, 100-112, 100-111, 100-110, 100-109, 100-108, 100-107, 100-106, 100-105, 100-104, 100-103, 100-102, or 100-101 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-20 mM KCl.
  • WSGR Docket No.47991-744.601 [0314] In some embodiments, the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-40, 0.1-39, 0.1-38, 0.1-37, 0.1-36, 0.1-35, 0.1-34, 0.1-33, 0.1-32, 0.1-31, 0.1-30, 0.1-29, 0.1-28, 0.1-27, 0.1-26, 0.1-25, 0.1-24, 0.1-23, 0.1-22, 0.1-21, 0.1-20, 0.1-19, 0.1-18, 0.1-17, 0.1-16, 0.1- 15, 0.1-14, 0.1-13, 0.1-12, 0.1-10, 0.1-9, 0.1-8, 0.1-7, 0.1-6, 0.1-5, 0.1-4, 0.1-3, 0.1-2, or 0.1-1 mM
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.2- 40, 0.3-40, 0.4-40, 0.5-40, 0.6-40, 0.7-40, 0.8-40, 0.9-40, 1-40, 2-40, 3-40, 4-40, 5-40, 6-40, 7-40, 8-40, 9-40, 10-40, 11-40, 12-40, 13-40, 14-40, 15-40, 16-40, 17-40, 18-40, 19-40, 20-40, 21-40, 22-40, 23-40, 24-40, 25-40, 26-40, 27-40, 28-40, 29-40, 30-40, 31-40, 32-40, 33-40, 34-40, 35-40, 36-40, 37-40, 38-40, or 39-40 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.5, 0.2-3.5, 0.3-3.5, 0.4-3.5, 0.5-3.5, 0.6-3.5, 0.7-3.5, 0.8-3.5, 0.9-3.5, 1.0-3.5, 1.1-3.5, 1.2-3.5, 1.3-3.5, 1.4-3.5, 1.5-3.5, 1.6-3.5, 1.7-3.5, 1.8-3.5, 1.9-3.5, 2.0-3.5, 2.1-3.5, 2.2-3.5, 2.3- 3.5, 2.4-3.5, 2.5-3.5, 2.6-3.5, 2.7-3.5, 2.8-3.5, 2.9-3.5, 3.0-3.5, 3.1-3.5, 3.2-3.5, 3.3-3.5, or 3.4-3.5 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.4, 0.1-3.3, 0.1-3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1- 2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50 mM Na 2 HPO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01-100, 0.02-100, 0.03-100, 0.04-100, 0.05-100, 0.06-100, 0.07-100, 0.08-100, 0.09-100, 0.1-100, 0.2-100, 0.3-100, 0.4-100, 0.5-100, 0.6-100, 0.7-100, 0.8-100, 0.9-100, 1-100, 2-100, 3-100, 4- 100, 5-100, 6-100, 7-100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45- 100, 50-100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01- 95, 0.01-90, 0.01-85, 0.01-80, 0.01-75, 0.01-70, 0.01-65, 0.01-60, 0.01-55, 0.01-50, 0.01-45, 0.01-40, 0.01-35, 0.01-30, 0.01-25, 0.01-20, 0.01-15, 0.01-10, 0.01-9, 0.01-8, 0.01-7, 0.01-6, 0.01-5, 0.01-4, 0.01- 3, 0.01-2, 0.01-1, 0.01-0.9, 0.01-0.8, 0.01-0.7, 0.01-0.6, 0.01-0.5, 0.01-0.4, 0.01-0.3, 0.01-0.2, 0.01-0.1, 0.01-0.09, 0.01-0.08, 0.01-0.07, 0.01-0.06, 0.01-0.05, 0.01-0.04, 0.01-0.03, or 0.01-0.02 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1- 3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, WSGR Docket No.47991-744.601 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1- 0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.0, 0.2-3.0, 0.3-3.0, 0.4-3.0, 0.5-3.0, 0.6-3.0, 0.7-3.0, 0.8-3.0, 0.9-3.0, 1.0-3.0, 1.2-3.0, 1.3-3.0, 1.4-3.0, 1.5-3.0, 1.6-3.0, 1.7-3.0, 1.8-3.0, 1.9-3.0, 2.0-3.0, 2.1- 3.0, 2.2-3.0, 2.3-3.0, 2.4-3.0, 2.5-3.0, 2.6-3.0, 2.7-3.0, 2.8-3.0, or 2.9-3.0 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM Na 2 HPO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01-100, 0.02-100, 0.03-100, 0.04-100, 0.05-100, 0.06-100, 0.07-100, 0.08-100, 0.09-100, 0.1-100, 0.2-100, 0.3-100, 0.4-100, 0.5-100, 0.6-100, 0.7-100, 0.8-100, 0.9-100, 1-100, 2-100, 3-100, 4- 100, 5-100, 6-100, 7-100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45- 100, 50-100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01- 95, 0.01-90, 0.01-85, 0.01-80, 0.01-75, 0.01-70, 0.01-65, 0.01-60, 0.01-55, 0.01-50, 0.01-45, 0.01-40, 0.01-35, 0.01-30, 0.01-25, 0.01-20, 0.01-15, 0.01-10, 0.01-9, 0.01-8, 0.01-7, 0.01-6, 0.01-5, 0.01-4, 0.01- 3, 0.01-2, 0.01-1, 0.01-0.9, 0.01-0.8, 0.01-0.7, 0.01-0.6, 0.01-0.5, 0.01-0.4, 0.01-0.3, 0.01-0.2, 0.01-0.1, 0.01-0.09, 0.01-0.08, 0.01-0.07, 0.01-0.06, 0.01-0.05, 0.01-0.04, 0.01-0.03, or 0.01-0.02 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1- 3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1- 0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.0, 0.2-3.0, 0.3-3.0, 0.4-3.0, 0.5-3.0, 0.6-3.0, 0.7-3.0, 0.8-3.0, 0.9-3.0, 1.0-3.0, 1.2-3.0, 1.3-3.0, 1.4-3.0, 1.5-3.0, 1.6-3.0, 1.7-3.0, 1.8-3.0, 1.9-3.0, 2.0-3.0, 2.1- 3.0, 2.2-3.0, 2.3-3.0, 2.4-3.0, 2.5-3.0, 2.6-3.0, 2.7-3.0, 2.8-3.0, or 2.9-3.0 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, WSGR Docket No.47991-744.601 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50 mM CaCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50, 0.2-50, 0.3-50, 0.4-50, 0.5-50, 0.6-50, 0.7-50, 0.8-50, 0.9-50, 1.0-50, -50, 1.1-50, 1.2- 50, 1.3-50, 1.4-50, 1.5-50, 1.6-50, 1.7-50, 1.8-50, 1.9-50, 2.0-50, 2.1-50, 2.2-50, 2.3-50, 2.4-50, 2.5-50, 2.6-50, 2.7-50, 2.8-50, 2.9-50, 3.0-50, 3.1-50, 3.2-50, 3.3-50, 3.4-50, 3.5-50, 3.6-50, 3.7-50, 3.8-50, 3.9- 50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 10-50, 15-50, 20-50, 25-50, 130-50, 35-50, 40-50, or 45-50 mM CaCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-45, 0.1-40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-15, 0.1-10, 0.1-5, 0.1-4, 0.1-4.9, 0.1-4.8, 0.1-4.7, 0.1-4.6, 0.1-4.5, 0.1-4.4, 0.1-4.3, 0.1-4.2, 0.1-4.1, 0.1-4.0, 0.1-3.9, 0.1-3.8, 0.1-3.7, 0.1-3.6, 0.1- 3.5, 0.1-3.4, 0.1-3.3, 0.1-3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.8,
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl 2 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl 2 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl 2 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50 mM MgCl 2 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50, 0.2-50, 0.3-50, 0.4-50, 0.5-50, 0.6-50, 0.7-50, 0.8-50, 0.9-50, 1.0-50, -50, 1.1-50, 1.2- 50, 1.3-50, 1.4-50, 1.5-50, 1.6-50, 1.7-50, 1.8-50, 1.9-50, 2.0-50, 2.1-50, 2.2-50, 2.3-50, 2.4-50, 2.5-50, 2.6-50, 2.7-50, 2.8-50, 2.9-50, 3.0-50, 3.1-50, 3.2-50, 3.3-50, 3.4-50, 3.5-50, 3.6-50, 3.7-50, 3.8-50, 3.9- 50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 10-50, 15-50, 20-50, 25-50, 130-50, 35-50, 40-50, or 45-50 mM MgCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-45, 0.1-40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-15, 0.1-10, 0.1-5, 0.1-4, 0.1-4.9, 0.1-4.8, 0.1-4.7, 0.1-4.6, 0.1-4.5, 0.1-4.4, 0.1-4.3, 0.1-4.2, 0.1-4.1, 0.1-4.0, 0.1-3.9, 0.1-3.8, 0.1-3.7, 0.1-3.6, 0.1- 3.5, 0.1-3.4, 0.1-3.3, 0.1-3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.8,
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, WSGR Docket No.47991-744.601 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl2.
  • the ASO is solubilized or diluted in a buffer further comprising 1-100 mM NaHCO3, 1-100 mM KHCO3, or a combination thereof.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100 mM NaHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-99, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM NaHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-100, 3-100, 4-100, 5-100, 6-100, 7- 100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60- 100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM NaHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.0-28.0, 24.0-27.9, 24.0- 27.8, 24.0-27.7, 24.0-27.6, 24.0-27.5, 24.0-27.4, 24.0-27.3, 24.0-27.2, 24.0-27.1, 24.0-27.0, 24.0-26.9, 24.0-26.8, 24.0-26.7, 24.0-26.6, 24.0-26.5, 24.0-26.4, 24.0-26.3, 24.0-26.2, 24.0-26.1, 24.0-26.0, 24.0- 25.9, 4.0-25.8, 24.0-25.7, 24.0-25.6, 24.0-25.5, 24.0-25.4, 24.0-25.3, 24.0-25.2, 24.0-25.1, 24.0-25.0, 24.0-24.9, 24.0-24.8, 24.0-24.7, 24.0-24.6, 24.0-24.5, 24.0-24.4, 24.0-24.3, 24.0-24.2, or 24.0-24.1 mM NaHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.1-28.0, 24.2-28.0, 24.3-28.0, 24.4-28.0, 24.5-28.0, 24.6-28.0, 24.7-28.0, 24.8-28.0, 24.9- 28.0, 25.0-28.0, 25.1-28.0, 25.2-28.0, 25.3-28.0, 25.4-28.0, 25.5-28.0, 25.6-28.0, 25.7-28.0, 25.8-28.0, 25.9-28.0, 26.0-28.0, 26.1-28.0, 26.2-28.0, 26.3-28.0, 26.4-28.0, 26.5-28.0, 26.6-28.0, 26.7-28.0, 26.8- 28.0, 26.9-28.0, 27.0-28.0, 27.1-28.0, 27.2-28.0, 27.3-28.0, 27.4-28.0, 27.5-28.0, 27.6-28.0, 27.7-28.0, 27.8-28.0, or 27.9-28.0 mM NaHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.0, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO3.
  • a buffer comprising 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1,
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100 mM KHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-99, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM KHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-100, 3-100, 4-100, 5-100, 6-100, 7- 100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60- 100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM KHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.0-28.0, 24.0-27.9, 24.0-27.8, 24.0-27.7, 24.0-27.6, 24.0-27.5, 24.0-27.4, 24.0-27.3, 24.0-27.2, 24.0-27.1, 24.0-27.0, 24.0-26.9, 24.0- 26.8, 24.0-26.7, 24.0-26.6, 24.0-26.5, 24.0-26.4, 24.0-26.3, 24.0-26.2, 24.0-26.1, 24.0-26.0, 24.0-25.9, 4.0-25.8, 24.0-25.7, 24.0-25.6, 24.0-25.5, 24.0-25.4, 24.0-25.3, 24.0-25.2, 24.0-25.1, 24.0-25.0, 24.0- 24.9, 24.0-24.8, 24.0-24.7, 24.0-24.6, 24.0-24.5, 24.0-24.4, 24.0-24.3, 24.0-24.2, or 24.0-24.1 mM KHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.1-28.0, 24.2-28.0, 24.3-28.0, 24.4-28.0, 24.5-28.0, 24.6-28.0, 24.7-28.0, 24.8-28.0, 24.9- 28.0, 25.0-28.0, 25.1-28.0, 25.2-28.0, 25.3-28.0, 25.4-28.0, 25.5-28.0, 25.6-28.0, 25.7-28.0, 25.8-28.0, 25.9-28.0, 26.0-28.0, 26.1-28.0, 26.2-28.0, 26.3-28.0, 26.4-28.0, 26.5-28.0, 26.6-28.0, 26.7-28.0, 26.8- 28.0, 26.9-28.0, 27.0-28.0, 27.1-28.0, 27.2-28.0, 27.3-28.0, 27.4-28.0, 27.5-28.0, 27.6-28.0, 27.7-28.0, 27.8-28.0, or 27.9-28.0 mM KHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.0, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM KHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM KHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM KHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-50 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-100, 0.01-100, 0.02-100, 0.03-100, 0.04-100, 0.05-100, 0.06-100, 0.07-100, 0.08-100, 0.09- 100, 0.1-100, 0.2-100, 0.3-100, 0.4-100, 0.5-100, 0.6-100, 0.7-100, 0.8-100, 0.9-100, 1-100, 2-100, 3- 100, 4-100, 5-100, 6-100, 7-100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 85-
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-95, 0-90, 0-85, 0-80, 0-75, 0-70, 0-65, 0-60, 0-55, 0-50, 0-45, 0-40, 0-35, 0-30, 0-25, 0-20, WSGR Docket No.47991-744.601 0-15, 0-10, 0-9, 0-8, 0-7, 0-6, 0-5, 0-4, 0-3, 0-2, 0-1, 0-0.9, 0-0.8, 0-0.7, 0-0.6, 0-0.5, 0-0.4, 0-0.3, 0-0.2, 0-0.1, 0-0.09, 0-0.08, 0-0.07, 0-0.06, 0-0.05, 0-0.04, 0-0.03, or 0-0.02 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01-95, 0.01- 90, 0.01-85, 0.01-80, 0.01-75, 0.01-70, 0.01-65, 0.01-60, 0.01-55, 0.01-50, 0.01-45, 0.01-40, 0.01-35, 0.01-30, 0.01-25, 0.01-20, 0.01-15, 0.01-10, 0.01-9, 0.01-8, 0.01-7, 0.01-6, 0.01-5, 0.01-4, 0.01-3, 0.01-2, 0.01-1, 0.01-0.9, 0.01-0.8, 0.01-0.7, 0.01-0.6, 0.01-0.5, 0.01-0.4, 0.01-0.3, 0.01-0.2, 0.01-0.1, 0.01-0.09, 0.01-0.08, 0.01-0.07, 0.01-0.06, 0.01-0.05, 0.01-0.04, 0.01-0.03, or 0.01-0.02 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-3.0, 0-2.9, 0-2.8, 0-2.7, 0-2.6, 0-2.5, 0-2.4, 0-2.3, 0-2.2, 0-2.1, 0-2.0, 0-1.9, 0-1.8, 0-1.7, 0-1.6, 0-1.5, 0-1.4, 0-1.3, 0- 1.2, 0-1.1, 0-1.0, 0-0.9, 0-0.8, 0-0.7, 0-0.6, 0-0.5, 0-0.4, 0-0.3, or 0-0.2 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.0, 0.1- 2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1- 0.4, 0.1-0.3, or 0.1-0.2 mM KH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-3.0, 0.1-3.0, 0.2-3.0, 0.3-3.0, 0.4-3.0, 0.5-3.0, 0.6-3.0, 0.7-3.0, 0.8-3.0, 0.9-3.0, 1.0-3.0, 1.2-3.0, 1.3-3.0, 1.4-3.0, 1.5-3.0, 1.6-3.0, 1.7-3.0, 1.8-3.0, 1.9-3.0, 2.0-3.0, 2.1-3.0, 2.2- 3.0, 2.3-3.0, 2.4-3.0, 2.5-3.0, 2.6-3.0, 2.7-3.0, 2.8-3.0, or 2.9-3.0 mM KH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM KH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM KH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM KH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-50 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-50, 0-45, 0-40, 0-35, 0-30, 0-25, 0-20, 0-19, 0-18, 0-17, 0-16, 0-15, 0-14, 0-13, 0-12, 0-11, 0-10, 0-9, 0-8, 0-7, 0-6, 0-5, 0-4, 0-3, 0-2, 0-1, 0-0.9, 0-0.8, 0-0.7, 0-0.6, 0-0.5, 0-0.4, 0-0.3, or 0-0.2 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50, 0.1-45, 0.1-40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-19, 0.1-18, 0.1-17, 0.1-16, 0.1-15, 0.1-14, 0.1-13, 0.1-12, 0.1-11, 0.1-10, 0.1-9, 0.1-8, 0.1-7, 0.1-6, 0.1-5, 0.1-4, 0.1-3, 0.1-2, 0.1-1, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-50, 0.1-50, 0.2-50, 0.3-50, 0.4- 50, 0.5-50, 0.6-50, 0.7-50, 0.8-50, 0.9-50, 1-50, 2-50, 3-50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 1-50, 11- 50, 12-50, 13-50, 14-50, 15-50, 16-50, 17-50, 18-50, 19-50, 20-50, 25-50, 30-50, 35-50, 40-50, or 45-50 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer WSGR Docket No.47991-744.601 comprising 0-20, 0.1-20, 0.2-20, 0.3-20, 0.4-20, 0.5-20, 0.6-20, 0.7-20, 0.8-20, 0.9-20, 1-20, 2-20, 3-20, 4-20, 5-20, 6-20, 7-20, 8-20, 9-20, 10-20, 11-20, 12-20, 13-20, 14-20, 15-20, 16-20, 17-20, 18-20, or 19- 20 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM NaH2PO4. In some embodiments, the ASO as described herein is solubilized or diluted in a buffer comprising at most 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM NaH2PO4.
  • the ASO is solubilized in the liquid composition that is not buffered by a buffering agent.
  • a buffering agent in a liquid formulation described herein is an agent other than the active pharmaceutical ingredient (e.g., an ASO described herein) in the formulation that buffers the pH of the formulation.
  • the ASO is solubilized in the liquid composition that comprises a buffering agent.
  • the buffering agent has a pKa at 25 °C of about 4.75; about 5.64; about 1.70, about 6.04 and about 9.09; about 3.1, about 4.7 and about 6.4; or about 6.50.
  • the buffering agent is an effective buffer at a pH range of from about 3.6 to 5.6, from about 5.5 to 6.5, from about 5.5 to 7.4, from about 3.0 to 6.2, or from about 5.8 to 7.2.
  • the buffering agent is selected from the group consisting of acetate, succinate, histidine, citrate, 2-[Bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol (Bis-Tris), and any combination thereof.
  • the ASO is stable in the pharmaceutical formulation for at least 1 or 2 or 3 years at -20 °C. In some embodiments, the ASO is stable in the pharmaceutical formulation for at least 1 or 2 or 3 years at 4 °C. In some embodiments, the ASO is stable in the pharmaceutical formulation for at least 1 or 2 or 3 years at 25 °C.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at -20 °C for at least 12 months.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at -20 °C for at least 36 months. In some embodiments, the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the WSGR Docket No.47991-744.601 pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is stable in the pharmaceutical WSGR Docket No.47991-744.601 formulation according to identification of a molecule with a molecular weight of 7197.2 ⁇ 4.0 Da according to LC/MS.
  • the pharmaceutical formulation has a shelf life of at least 1 or 2 or 3 years when stored at -20 °C. In some embodiments, the pharmaceutical formulation has a shelf life of at least 1 or 2 or 3 years when stored at 4 °C. In some embodiments, the pharmaceutical formulation has a shelf life of at least 1 or 2 or 3 years when stored at 25 °C.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at -20 °C.
  • the ASO is stable in the pharmaceutical formulation after storage of the pharmaceutical formulation at -20 °C for at least 12 months.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 4 °C.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 25 °C.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 30 °C.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 37 °C.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 40 °C.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, WSGR Docket No.47991-744.601 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity.
  • the pharmaceutical formulation has a shelf life of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years when stored at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at -20 °C for at least 12 months. In some embodiments, the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at -20 °C for at least 36 months.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC WSGR Docket No.47991-744.601 relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.92 is not more than 2.5% after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, WSGR Docket No.47991-744.601 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at -20 °C for at least 12 months. In some embodiments, the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at -20 °C for at least 36 months.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the WSGR Docket No.47991-744.601 pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent impurities in the pharmaceutical formulation according to an HPLC relative retention time of 0.98 is not more than 3.5% after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at -20 °C for at least 12 months. In some embodiments, the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at -20 °C for at least 36 months.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to WSGR Docket No.47991-744.601 HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of any unspecified impurities in the pharmaceutical WSGR Docket No.47991-744.601 formulation according to HPLC is not more than 1.8% after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at -20 °C for at least 12 months.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at -20 °C for at least 36 months.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the percent of total impurities in the pharmaceutical formulation according to HPLC is not more than 10% after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at -20 °C for at least 12 months.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at -20 °C for at least 36 months.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the pH of the pharmaceutical composition is from 6.6 to 7.6 after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, WSGR Docket No.47991-744.601 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at -20 °C for at least 12 months. In some embodiments, the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at -20 °C for at least 36 months.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from WSGR Docket No.47991-744.601 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the osmolality of the pharmaceutical composition is from 310 to 360 mOsm/kg after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at -20 °C for at least 12 months. In some embodiments, no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at -20 °C for at least 36 months.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, WSGR Docket No.47991-744.601 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation or no formation of particulate that has a dimension larger than 50 ⁇ m is observed after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, WSGR Docket No.47991-744.601 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at -20 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at -20 °C for at least 12 months.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at -20 °C for at least 24 months. In some embodiments, no observable particulate formation is observed after storage of the pharmaceutical formulation at -20 °C for at least 36 months.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 4 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 25 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 30 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 37 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 40 °C for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 4 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 25 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 WSGR Docket No.47991-744.601 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 30 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 37 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • no observable particulate formation is observed after storage of the pharmaceutical formulation at 40 °C at 55%, 60%, 65%, 70% or 75% relative humidity for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, or 52 weeks, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months, or 1, 2, 3, 4, or 5 years.
  • the ASO is solubilized in a buffer without Na 2 HPO 4 and/or NaH 2 PO 4 .
  • the ASO is solubilized or diluted in a buffer further comprising carbohydrates.
  • the carbohydrates comprise D-glucose.
  • the ASO is solubilized or diluted in a buffer further comprising 1-100 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-29, 1-28, 1-27, 1-26, 1-25, 1-24, 1-23, 1-22, 1-21, 1-20, 1-19, 1-18, 1-17, 1-16, 1-15, 1-14, 1-13, 1-12, 1-11, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-100, 3-100, 4-100, 5-100, 6-100, 7-100, 8-100, 9- 100, 10-100, 11-100, 12-100, 13-100, 14-100, 15-100, 16-100, 17-100, 18-100, 19-100, 20-100, 21-100, 22-100, 23-100, 24-100, 25-100, 26-100, 29-100, 28-100, 29-100, 30-100, 35-100, 40-100, 45-100, 50- 100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-30, 3- 30, 4-30, 5-30, 6-30, 7-30, 8-30, 9-30, 10-30, 11-30, 12-30, 13-30, 14-30, 15-30, 16-30, 17-30, 18-30, 19- 30, 20-30, 21-30, 22-30, 23-30, 24-30, 25-30, 26-30, 27-30, 28-30, or 29-30 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM D- glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM D-glucose. In some embodiments, the ASO as described herein is solubilized or diluted in a buffer comprising 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-29, 1-28, 1-27, 1-26, 1-25, 1-24, 1-23, 1-22, 1-21, 1-20, 1-19, 1-18, 1-17, 1-16, 1-15, 1-14, 1-13, 1-12, 1-11, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-100, 3-100, 4-100, 5-100, 6-100, 7-100, 8-100, 9- 100, 10-100, 11-100, 12-100, 13-100, 14-100, 15-100, 16-100, 17-100, 18-100, 19-100, 20-100, 21-100, 22-100, 23-100, 24-100, 25-100, 26-100, 29-100, 28-100, 29-100, 30-100, 35-100, 40-100, 45-100, 50- 100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-30, 3-30, 4- 30, 5-30, 6-30, 7-30, 8-30, 9-30, 10-30, 11-30, 12-30, 13-30, 14-30, 15-30, 16-30, 17-30, 18-30, 19-30, 20-30, 21-30, 22-30, 23-30, 24-30, 25-30, 26-30, 27-30, 28-30, or 29-30 mM glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM glucose. In some embodiments, the ASO as described herein is solubilized or diluted in a buffer comprising 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM glucose.
  • the ASO is solubilized or diluted in a buffer comprising 25-250 mM NaCl, 0.1-20 mM KCl, 0.1-50 mM Na 2 HPO 4 , 0.1-50 mM NaH 2 PO 4 , 0.1-50 mM CaCl 2 , and 0.1-50 mM MgCl 2 .
  • the ASO is solubilized or diluted in a buffer comprising 150 mM NaCl, 3.0 mM KCl, 0.7 mM Na 2 HPO 4 , 0.3 mM NaH 2 PO 4 , 0.79 mM MgCl 2 , and 1.4 mM CaCl 2 .
  • the ASO is solubilized or diluted in a buffer further comprising an antioxidant.
  • the antioxidant is t-butylhydroxyquinoline (TBHQ), butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), vitamin E, or any combination thereof.
  • the ASO is solubilized or diluted in a buffer further comprising an antioxidant, wherein the antioxidant is ascorbic acid (vitamin C), glutathione, lipoic acid, uric acid, carotenes, ⁇ -tocopherol (vitamin E), ubiquinol (coenzyme Q), or any combination thereof.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-250 mM NaCl, 0.1-20 mM KCl, 0-50 mM KH2PO4, 1-100 mM NaHCO3, 0-50 mM Na2HPO4, 1-100 mM D-glucose, 0.1-50 mM CaCl2, 0.1-50 mM MgCl2, or any combination thereof.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-250 mM NaCl, 0.1-20 mM KCl, 0-50 mM KH2PO4, 1-100 mM NaHCO3, 0-50 mM Na2HPO4, 1-100 mM D-glucose, 0.1-50 mM CaCl2, and 0.1-50 mM MgCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 127 mM NaCl, 1.0 mM KCl, 1.2 mM KH2PO4, 26 mM NaHCO3, 10 mM D-glucose, 2.4 mM CaCl2, and 1.3 mM MgCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 119 mM NaCl, 26.2 mM NaHCO3, 2.5 mM KCl, 1 mM NaH2PO4, 1.3 mM MgCl2, 10 mM glucose, and 2.5 mM CaCl2.
  • the pharmaceutical composition does not comprise a preservative.
  • the pharmaceutical composition comprises a preservative.
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 5-250 mg/mL.
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 5-250, 5-247.5, 5-245, 5-242.5, 5-240, 5-237.5, 5-235, 5-232.5, 5-230, 5- 227.5, 5-225, 5-225.5, 5-220, 5-217.5, 5-215, 5-212.5, 5-210, 5-205.5, 5-205, 5-202.5, 5-200, 5-197.5, 5- 195, 5-192.5, 5-190, 5-187.5, 5-185, 5-182.5, 5-180, 5-177.5, 5-175, 5-172.5, 5-170, 5-167.5, 5-165, 5- 162.5, 5-160, 5-157.5, 5-155, 5-152.5, 5-150, 5-147.5, 5-145, 5-142.5, 5-140, 5-137.5, 5-135, 5-132.5, 5- 130, 5-127.5, 5-125, 5-122.5, 5-120, 5-117.5, 5-115, 5-112.5, 5-110, 5-107.5, 5-105, 5-102.5, 5-100, 5- 97.5, 5-95, 5-9
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 10-250, 15-250, 20-250, 25-250, 30-250, 35-250, 40-250, 45-250, 50-250, 55-250, 60-250, 65-250, 70-250, 75-250, 80-250, 85-250, 90-250, 95- 250, 100-250, 105-250, 110-250, 115-250, 120-250, 125-250, 130-250, 135-250, 140-250, 145-250, 150- 250, 155-250, 160-250, 165-250, 170-250, 175-250, 180-250, 185-250, 190-250, or 195-250, 200-250, 205-250, 210-250, 215-250, 220-250, 225-250, 230-250, 235-250, 240-250, or 245-250 mg/mL.
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110,
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110,
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 0.1 mg/mL to 250 mg/mL. In some embodiments, the ASO as described herein is present in the pharmaceutical composition at a concentration of about 0.1 mg/mL, 0.5 mg/mL, 1 mg/mL, 2 mg/mL, 2.5 mg/mL, 3 mg/mL, 4 mg/mL, 5 mg/mL, 6 mg/mL, 7 mg/mL, 8 mg/mL, 9 mg/mL, 10 mg/mL, 11 mg/mL, 12 mg/mL, 13 mg/mL, 14 mg/mL, 15 mg/mL, 16 mg/mL, 17 mg/mL, 18 mg/mL, 19 mg/mL, or 20 mg/mL.
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of about 30 mg/mL, 40 mg/mL, 50 mg/mL, 60 mg/mL, 70 mg/mL, 80 mg/mL, 90 mg/mL, 100 mg/mL, 110 mg/mL, 120 mg/mL, 130 mg/mL, 140 mg/mL, 150 mg/mL, 160 mg/mL, 170 mg/mL, 180 mg/mL, 190 mg/mL, or 200 mg/mL.
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of about 11 mg/mL, 22 mg/mL, 33 mg/mL, 44 mg/mL, 55 mg/mL, 66 mg/mL, 77 mg/mL, 88 mg/mL, 99 mg/mL, or 100 mg/mL in the diluent.
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 0.1 mg/mL to 250 mg/mL, from 0.2 mg/mL to 250 mg/mL, from 0.3 mg/mL to 250 mg/mL, from 0.4 mg/mL to 250 mg/mL, from 0.5 mg/mL to 250 mg/mL, from 0.6 mg/mL to 250 mg/mL, from 0.7 mg/mL to 250 mg/mL, from 0.8 mg/mL to 250 mg/mL, from 0.9 mg/mL to 250 mg/mL, from 1.0 mg/mL to 250 mg/mL, from 1.1 mg/mL to 250 mg/mL, from 1.2 mg/mL to 250 mg/mL, from 1.3 mg/mL to 250 mg/mL, from 1.4 mg/mL to 250 mg/mL, from 1.5 mg/mL to 250 mg/mL, from 1.6 mg/mL to 250 mg/mL, from 1.7 mg/
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 0.1 mg/mL to 250 mg/mL, from 0.1 mg/mL to 245 mg/mL, from 0.1 mg/mL to 240 mg/mL, from 0.1 mg/mL to 235 mg/mL, from 0.1 mg/mL to 230 mg/mL, from 0.1 mg/mL to 225 mg/mL, from 0.1 mg/mL to 220 mg/mL, from 0.1 mg/mL to 215 mg/mL, from 0.1 mg/mL to 210 mg/mL, from 0.1 mg/mL to 205 mg/mL, from 0.1 mg/mL to 200 mg/mL, from 0.1 mg/mL to 195 mg/mL, from 0.1 mg/mL to 190 mg/mL, from 0.1 mg/mL to 185 mg/mL, from 0.1 mg/mL to 180 mg/mL, from 0.1 mg/m/m
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59,
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60,
  • compositions comprising the agent, e.g., antisense oligonucleotide or antisense oligomer, of the described compositions and for use in any of the described methods can be prepared according to conventional techniques well-known in the pharmaceutical industry and described in the published literature.
  • a pharmaceutical composition for treating a subject comprises an effective amount of any antisense oligomer as described herein, or a pharmaceutically acceptable salt, solvate, hydrate or ester thereof.
  • the pharmaceutical composition described herein further comprises a pharmaceutically acceptable excipient, carrier, or diluent.
  • compositions can be formulated in a conventional manner using one or more pharmaceutically acceptable inactive ingredients that facilitate processing of the active compounds into preparations that can be used pharmaceutically.
  • a proper formulation is dependent upon the route of administration chosen and a summary of pharmaceutical compositions can be found, for example, in Remington: The Science and Practice of Pharmacy, Nineteenth Ed (Easton, Pa.: Mack Publishing Company, 1995); Hoover, John E., Remington’s Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania 1975; Liberman, H.A. and Lachman, L., Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, N.Y., 1980; and Pharmaceutical Dosage Forms and Drug Delivery Systems, Seventh Ed.
  • the pharmaceutical composition facilitates administration of the compound to an organism.
  • Such compositions may comprise buffers such as neutral buffered saline, phosphate buffered saline and the like; carbohydrates such as glucose, mannose, sucrose or dextrans, mannitol; proteins; WSGR Docket No.47991-744.601 polypeptides or amino acids such as glycine; antioxidants; chelating agents such as EDTA or glutathione; adjuvants (e.g., aluminum hydroxide); and preservatives.
  • composition and “pharmaceutical formulation” (or “formulation”) are used interchangeably and denote a mixture or solution comprising a therapeutically effective amount of an active pharmaceutical ingredient together with one or more pharmaceutically acceptable excipients to be administered to a subject, e.g., a human in need thereof.
  • pharmaceutically acceptable denotes an attribute of a material which is useful in preparing a pharmaceutical composition that is generally safe, non-toxic, and neither biologically nor otherwise undesirable and is acceptable for veterinary as well as human pharmaceutical use.
  • “Pharmaceutically acceptable” can refer a material, such as a carrier or diluent, which does not abrogate the biological activity or properties of the compound, and is relatively nontoxic, i.e., the material may be administered to an individual without causing undesirable biological effects or interacting in a deleterious manner with any of the components of the composition in which it is contained.
  • compositions are prepared with carriers that will protect the components of the composition against rapid elimination from the body, such as a controlled release formulation, including implants and microencapsulated delivery systems.
  • Biodegradable, biocompatible polymers can be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid. Methods for preparation of such formulations will be apparent to those skilled in the art. The materials can also be obtained commercially from Alza Corporation and Nova Pharmaceuticals, Inc. Liposomal suspensions (including liposomes targeted to infected cells with monoclonal antibodies to viral anti-gens) can also be used as pharmaceutically acceptable carriers. These can be prepared according to methods known to those skilled in the art, for example, as described in U.S. Pat. No.4,522,811, of which entire content is incorporated herein by reference.
  • compositions or formulations comprising the agent, e.g., antisense oligonucleotide, of the described compositions and for use in any of the described methods can be prepared according to conventional techniques well-known in the pharmaceutical industry and described in the published literature.
  • a pharmaceutical composition or formulation for treating a subject comprises an effective amount of any antisense oligomer as described herein, or a pharmaceutically acceptable salt, solvate, hydrate or ester thereof.
  • the pharmaceutical formulation comprising an antisense oligomer may further comprise a pharmaceutically acceptable excipient, diluent or carrier.
  • salts are suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response, etc., and are commensurate with a reasonable benefit/risk ratio. (See, e.g., S. M. Berge, et al., J. Pharmaceutical Sciences, 66: 1-19 (1977), incorporated herein by reference for this purpose.
  • the salts can be prepared in situ during the final isolation and purification of the compounds, or separately by reacting the free base function with a suitable organic acid.
  • Examples of pharmaceutically acceptable, nontoxic acid addition salts are salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other documented methodologies such as ion exchange.
  • inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid
  • organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other documented methodologies such as ion exchange.
  • salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2- hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate
  • Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like.
  • Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, lower alkyl sulfonate and aryl sulfonate.
  • a pharmaceutical formulation comprising: an antisense oligomer (ASO as described herein), wherein the ASO as described herein comprises a sequence with at least 80% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099; and a pharmaceutically acceptable diluent; wherein about 0.1, 0.5, 1, 2.5, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 22.5, 25, 27.5, 30, 32.5, 35, 37.5, 40, 42.5, 45, 47.5, 50, 52.5, 55, 57.5, 60, 62.5, 65, 67.5, 70, 72.5, 75, 77.5, 80, 82.5, 85, 87.5, 90, 92.5, 95, 97.5, 100, 102.5, 105, 107.5, 110, 112.5, 115, 117.5, 120, 122.5, 125, 127.5, 130, 132.5, 135, 137.5, 140, 142.5, 145, 147.5, 150, 15
  • the ASO as described herein comprises a sequence with at least 80% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • WSGR Docket No.47991-744.601 [0374]
  • the ASO as described herein comprises a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304- 1099.
  • the ASO as described herein consists of a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the ASO as described herein consists of a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the ASO as described herein is a compound having the structure depicted in formula (I) (free acid): (I), or a salt thereof.
  • the ASO as described herein is a compound having the structure depicted in formula (II) (sodium salt): WSGR Docket No.47991-744.601 (II).
  • an ASO described herein is all-P-ambo-2'-O-(2-methoxyethyl)-P-thioadenylyl- (3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-5-methyl-P-thiouridylyl- (3' ⁇ 5')-2'-O-(2-methoxyethyl)-5-methyl-P-thiouridylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl- (3' ⁇ 5')-2'-O-(2-methoxyethyl)-2'-O-(2-methoxyethyl)-P-thioguanylyl- (3' ⁇ 5')-2'-O-(2-methoxyethyl)-2'-O-(2-methoxyethyl
  • an ASO described herein is a sodium salt of all-P-ambo-2'-O-(2-methoxyethyl)- P-thioadenylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-5-methyl- P-thiouridylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-5-methyl-P-thiouridylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)- P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-
  • the ASO as described herein is dissolved or suspended in a solution at a concentration of from 5-250, 5-247.5, 5-245, 5-242.5, 5-240, 5-237.5, 5-235, 5-232.5, 5-230, 5-227.5, 5- 225, 5-225.5, 5-220, 5-217.5, 5-215, 5-212.5, 5-210, 5-205.5, 5-205, 5-202.5, 5-200, 5-197.5, 5-195, 5- 192.5, 5-190, 5-187.5, 5-185, 5-182.5, 5-180, 5-177.5, 5-175, 5-172.5, 5-170, 5-167.5, 5-165, 5-162.5, 5- 160, 5-157.5, 5-155, 5-152.5, 5-150, 5-147.5, 5-145, 5-142.5, 5-140, 5-137.5, 5-135, 5-132.5, 5-130, 5- 127.5, 5-125, 5-122.5, 5-120, 5-117.5, 5-115, 5-112.5, 5-110, 5-107.5, 5-105, 5-102.5, 5-100, 5-97.5, 5- 95
  • the ASO as described herein is dissolved or suspended in a solution at a concentration of from 10-250, 15-250, 20-250, 25-250, 30-250, 35-250, 40- 250, 45-250, 50-250, 55-250, 60-250, 65-250, 70-250, 75-250, 80-250, 85-250, 90-250, 95-250, 100-250, 105-250, 110-250, 115-250, 120-250, 125-250, 130-250, 135-250, 140-250, 145-250, 150-250, 155-250, 160-250, 165-250, 170-250, 175-250, 180-250, 185-250, 190-250, or 195-250, 200-250, 205-250, 210- 250, 215-250, 220-250, 225-250, 230-250, 235-250, 240-250, or 245-250 mg/mL.
  • the ASO as described herein is dissolved or suspended in a solution at a concentration of from at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109
  • the ASO as described herein is dissolved or suspended in a solution at a concentration of from at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, WSGR Docket No.47991-744.601 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104
  • the ASO as described herein is dissolved or suspended in a solution at a concentration of from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110
  • the pharmaceutically acceptable diluent comprises an artificial cerebral spinal fluid (aCSF) solution.
  • the solution comprises a cerebral spinal fluid (CSF) sample from the subject.
  • the ASO as described herein is solubilized or diluted in an iso-tonic solution.
  • the ASO as described herein is solubilized or diluted in a phosphate- buffered (pH 6.6-7.6) solution.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered (pH 6.0-8.0) solution.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered (pH 5.0-8.0) solution. In some embodiments, the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 4.5-8.5, pH 4.6-8.5, pH 4.7-8.5, pH 4.8-8.5, pH 4.9-8.5, pH5.0-8.5, pH 5.1-8.5, pH5.2-8.5, pH 5.3-8.5, pH5.4-8.5, pH 5.5-8.5, pH5.6-8.5, pH 5.7-8.5, pH 5.8-8.5, H 5.9-8.5, pH 6.0-8.5, pH 6.1-8.5, pH 6.2-8.5, pH 6.3-8.5, pH 6.4-8.5, pH 6.5-8.5, pH 6.6-8.5, pH 6.7-8.5, pH 6.8-8.5, pH 6.9-8.5, pH 7.0-8.5, pH 7.1-8.5, pH 7.2-8.5, pH 7.3-8.5, pH 7.4-8.5, pH 7.5-8.5, pH
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 4.5-8.3, pH 4.5-8.2, pH 4.5-8.1, pH 4.5- 8.0, pH 4.5-7.9, pH 4.5-7.8, pH 4.5-7.7, pH 4.5-7.6, pH 4.5-7.5, pH 4.5-7.4, pH 4.5-7.3, pH 4.5-7.2, pH 4.5-7.1, pH 4.5-7.0, pH 4.5-6.9, pH 4.5-6.8, pH 4.5-6.7, pH 4.5-6.6, pH 4.5-6.5, pH 4.5-6.4, pH 4.5-6.3, pH 4.5-6.2, pH 4.5-6.1, pH 4.5-6.0, pH 4.5-5.9, pH 4.5-5.8, pH 4.5-5.7, pH 4.5-5.6, pH 4.5-5.5, pH 4.5- WSGR Docket No.47991-744.601 5.4, pH 4.5-5.3, pH 4.5-5.2, pH 4.5-5.1, pH 4.5-5.0, pH 4.5-4.9
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-7.6, pH 6.1-7.6, pH 6.2-7.6, pH 6.3-7.6, pH 6.4-7.6, pH 6.5-7.6, pH 6.6-7.6, pH 6.7- 7.6, pH 6.8-7.6, pH 6.9-7.6, pH 7.0-7.6, pH 7.1-7.6, pH 7.2-7.6, pH 7.3-7.6, pH 7.4-7.6, or pH 7.5-7.6.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.6-8.0, pH 6.6-7.9, pH 6.6-7.8, pH 6.6-7.7, pH 6.6-7.6, pH 6.6-7.5, pH 6.6-7.4, pH 6.6- 7.3, pH 6.6-7.2, pH 6.6-7.1, pH 6.6-7.0, pH 6.6-6.9, pH 6.6-6.8, or pH 6.6-6.7.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-8.0, pH 6.1-8.0, pH 6.2-8.0, pH 6.3-8.0, pH 6.4-8.0, pH 6.5-8.0, pH 6.6-8.0, pH 6.7-8.0, pH 6.8-8.0, pH 6.9-8.0, pH 7.0-8.0, pH 7.1-8.0, pH 7.2-8.0, pH 7.3-8.0, pH 7.4-8.0, pH 7.5-8.0, pH 7.6-8.0, pH 7.7-8.0, pH 7.8- 8.0, or pH 7.9-8.0.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-7.9, pH 6.0-7.8, pH 6.0-7.7, pH 6.0-7.6, pH 6.0-7.5, pH 6.0-7.4, pH 6.0-7.3, pH 6.0-7.2, pH 6.0-7.1, pH 6.0-7.0, pH 6.0-6.9, pH 6.0-6.8, pH 6.0-6.7, pH 6.0-6.6, pH 6.0- 6.5, pH 6.0-6.4, pH 6.0-6.3, pH 6.0-6.2, or pH 6.0-6.1.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 5.7-8.5, 5.8-8.4, 5.9-8.3, 6.0-8.2, 6.1-8.1, 6.2-8.0, 6.3-7.9, 6.4-7.8, 6.5-7.7, or 6.6-7.6.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH about 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0.6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, or 8.0.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0.6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, or 8.0.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-250 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-250, 30-250, 35-250, 40-250, 45-250, 50-250, 55-250, 60-250, 65-250, 70-250, 75-250, 80-250, 85-250, 90-250, 95-250, 100-250, 105-250, 110-250, 115-250, 120-250, 125-250, 130-250, 135- 250, 140-250, 145-250, 150-250, 155-250, 160-250, 165-250, 170-250, 175-250, 180-250, 185-250, 190- 250, 195-250, 200-250, 205-250, 210-250, 215-250, 220-250, 225-250, 230-250, 235-250, 240-250, or 245-250 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-245, 25-240, 25-235, 25-230, 25-225, 25-220, 25-215, 25-210, 25-205, 25-200, 25- 195, 25-190, 25-185, 25-180, 25-175, 25-170, 25-165, 25-160, 25-155, 25-150, 25-145, 25-140, 25-135, 25-130, 25-125, 25-120, 25-115, 25-110, 25-105, 25-110, 25-105, 25-100, 25-95, 25-90, 25-85, 25-80, 25-75, 25-70, 25-65, 25-60, 25-55, 25-50, 25-45, 25-40, 25-35, or 25-30 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 30-245, 35- 240, 40-235, 45-230, 50-225, 55-220, 60-215, 65-210, 70-205, 75-200, 80-195, 85-190, 90-185, 95-180, 100-175, 105-170, 110-165, 115-160, 120-155, 125-150, 130-145 or 135-140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 100-140, 101- 140, 102-140, 103-140, 104-140, 105-140, 106-140, 107-140, 108-140, 109-140, 110-140, 111-140, 112- WSGR Docket No.47991-744.601 140, 113-140, 114-140, 115-140, 116-140, 117-140, 118-140, 119-140, 120-140, 121-140, 122-140, 123- 140, 124-140, 125-140, 126-140, 127-140, 128-140, 129-140, 130-140, 131-140, 132-140, 133-140, 134- 140, 135-140, 136-140, 137-140, 138-140, or 139-140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 100-139, 100-138, 100-137, 100-136, 100-135, 100-134, 100-133, 100-132, 100-131, 100-130, 100-129, 100-128, 100-127, 100-126, 100-125, 100-124, 100-123, 100-122, 100-121, 100-120, 100-119, 100-118, 100-117, 100-116, 100-115, 100-114, 100-113, 100-112, 100-111, 100-110, 100-109, 100-108, 100-107, 100-106, 100-105, 100-104, 100-103, 100-102, or 100-101 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-20 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-40, 0.1-39, 0.1-38, 0.1-37, 0.1-36, 0.1-35, 0.1-34, 0.1-33, 0.1-32, 0.1-31, 0.1-30, 0.1-29, 0.1-28, 0.1-27, 0.1-26, 0.1-25, 0.1-24, 0.1-23, 0.1-22, 0.1-21, 0.1-20, 0.1-19, 0.1-18, 0.1-17, 0.1-16, 0.1- 15, 0.1-14, 0.1-13, 0.1-12, 0.1-10, 0.1-9, 0.1-8, 0.1-7, 0.1-6, 0.1-5, 0.1-4, 0.1-3, 0.1-2, or 0.1-1 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.2- 40, 0.3-40, 0.4-40, 0.5-40, 0.6-40, 0.7-40, 0.8-40, 0.9-40, 1-40, 2-40, 3-40, 4-40, 5-40, 6-40, 7-40, 8-40, 9-40, 10-40, 11-40, 12-40, 13-40, 14-40, 15-40, 16-40, 17-40, 18-40, 19-40, 20-40, 21-40, 22-40, 23-40, 24-40, 25-40, 26-40, 27-40, 28-40, 29-40, 30-40, 31-40, 32-40, 33-40, 34-40, 35-40, 36-40, 37-40, 38-40, or 39-40 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.5, 0.2-3.5, 0.3-3.5, 0.4-3.5, 0.5-3.5, 0.6-3.5, 0.7-3.5, 0.8-3.5, 0.9-3.5, 1.0-3.5, 1.1-3.5, 1.2-3.5, 1.3-3.5, 1.4-3.5, 1.5-3.5, 1.6-3.5, 1.7-3.5, 1.8-3.5, 1.9-3.5, 2.0-3.5, 2.1-3.5, 2.2-3.5, 2.3- 3.5, 2.4-3.5, 2.5-3.5, 2.6-3.5, 2.7-3.5, 2.8-3.5, 2.9-3.5, 3.0-3.5, 3.1-3.5, 3.2-3.5, 3.3-3.5, or 3.4-3.5 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.4, 0.1-3.3, 0.1-3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1- 2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, WSGR Docket No.47991-744.601 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01-100, 0.02-100, 0.03-100, 0.04-100, 0.05-100, 0.06-100, 0.07-100, 0.08-100, 0.09-100, 0.1-100, 0.2-100, 0.3-100, 0.4-100, 0.5-100, 0.6-100, 0.7-100, 0.8-100, 0.9-100, 1-100, 2-100, 3-100, 4- 100, 5-100, 6-100, 7-100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45- 100, 50-100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM Na 2 HPO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01- 95, 0.01-90, 0.01-85, 0.01-80, 0.01-75, 0.01-70, 0.01-65, 0.01-60, 0.01-55, 0.01-50, 0.01-45, 0.01-40, 0.01-35, 0.01-30, 0.01-25, 0.01-20, 0.01-15, 0.01-10, 0.01-9, 0.01-8, 0.01-7, 0.01-6, 0.01-5, 0.01-4, 0.01- 3, 0.01-2, 0.01-1, 0.01-0.9, 0.01-0.8, 0.01-0.7, 0.01-0.6, 0.01-0.5, 0.01-0.4, 0.01-0.3, 0.01-0.2, 0.01-0.1, 0.01-0.09, 0.01-0.08, 0.01-0.07, 0.01-0.06, 0.01-0.05, 0.01-0.04, 0.01-0.03, or 0.01-0.02 mM Na 2 HPO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1- 3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1- 0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM Na 2 HPO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.0, 0.2-3.0, 0.3-3.0, 0.4-3.0, 0.5-3.0, 0.6-3.0, 0.7-3.0, 0.8-3.0, 0.9-3.0, 1.0-3.0, 1.2-3.0, 1.3-3.0, 1.4-3.0, 1.5-3.0, 1.6-3.0, 1.7-3.0, 1.8-3.0, 1.9-3.0, 2.0-3.0, 2.1- 3.0, 2.2-3.0, 2.3-3.0, 2.4-3.0, 2.5-3.0, 2.6-3.0, 2.7-3.0, 2.8-3.0, or 2.9-3.0 mM Na 2 HPO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM Na 2 HPO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM Na2HPO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01-100, 0.02-100, 0.03-100, 0.04-100, 0.05-100, 0.06-100, 0.07-100, 0.08-100, 0.09-100, 0.1-100, 0.2-100, 0.3-100, 0.4-100, 0.5-100, 0.6-100, 0.7-100, 0.8-100, 0.9-100, 1-100, 2-100, 3-100, 4- WSGR Docket No.47991-744.601 100, 5-100, 6-100, 7-100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45- 100, 50-100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01- 95, 0.01-90, 0.01-85, 0.01-80, 0.01-75, 0.01-70, 0.01-65, 0.01-60, 0.01-55, 0.01-50, 0.01-45, 0.01-40, 0.01-35, 0.01-30, 0.01-25, 0.01-20, 0.01-15, 0.01-10, 0.01-9, 0.01-8, 0.01-7, 0.01-6, 0.01-5, 0.01-4, 0.01- 3, 0.01-2, 0.01-1, 0.01-0.9, 0.01-0.8, 0.01-0.7, 0.01-0.6, 0.01-0.5, 0.01-0.4, 0.01-0.3, 0.01-0.2, 0.01-0.1, 0.01-0.09, 0.01-0.08, 0.01-0.07, 0.01-0.06, 0.01-0.05, 0.01-0.04, 0.01-0.03, or 0.01-0.02 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1- 3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1- 0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.0, 0.2-3.0, 0.3-3.0, 0.4-3.0, 0.5-3.0, 0.6-3.0, 0.7-3.0, 0.8-3.0, 0.9-3.0, 1.0-3.0, 1.2-3.0, 1.3-3.0, 1.4-3.0, 1.5-3.0, 1.6-3.0, 1.7-3.0, 1.8-3.0, 1.9-3.0, 2.0-3.0, 2.1- 3.0, 2.2-3.0, 2.3-3.0, 2.4-3.0, 2.5-3.0, 2.6-3.0, 2.7-3.0, 2.8-3.0, or 2.9-3.0 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50 mM CaCl 2 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50, 0.2-50, 0.3-50, 0.4-50, 0.5-50, 0.6-50, 0.7-50, 0.8-50, 0.9-50, 1.0-50, -50, 1.1-50, 1.2- 50, 1.3-50, 1.4-50, 1.5-50, 1.6-50, 1.7-50, 1.8-50, 1.9-50, 2.0-50, 2.1-50, 2.2-50, 2.3-50, 2.4-50, 2.5-50, 2.6-50, 2.7-50, 2.8-50, 2.9-50, 3.0-50, 3.1-50, 3.2-50, 3.3-50, 3.4-50, 3.5-50, 3.6-50, 3.7-50, 3.8-50, 3.9- 50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 10-50, 15-50, 20-50, 25-50, 130-50, 35-50, 40-50, or 45-50 mM CaCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-45, 0.1-40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-15, 0.1-10, 0.1-5, 0.1-4, 0.1-4.9, 0.1-4.8, 0.1-4.7, 0.1-4.6, 0.1-4.5, 0.1-4.4, 0.1-4.3, 0.1-4.2, 0.1-4.1, 0.1-4.0, 0.1-3.9, 0.1-3.8, 0.1-3.7, 0.1-3.6, 0.1- 3.5, 0.1-3.4, 0.1-3.3, 0.1-3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.8,
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl2.
  • the ASO as described herein is WSGR Docket No.47991-744.601 solubilized or diluted in a buffer comprising at most 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50 mM MgCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50, 0.2-50, 0.3-50, 0.4-50, 0.5-50, 0.6-50, 0.7-50, 0.8-50, 0.9-50, 1.0-50, -50, 1.1-50, 1.2- 50, 1.3-50, 1.4-50, 1.5-50, 1.6-50, 1.7-50, 1.8-50, 1.9-50, 2.0-50, 2.1-50, 2.2-50, 2.3-50, 2.4-50, 2.5-50, 2.6-50, 2.7-50, 2.8-50, 2.9-50, 3.0-50, 3.1-50, 3.2-50, 3.3-50, 3.4-50, 3.5-50, 3.6-50, 3.7-50, 3.8-50, 3.9- 50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 10-50, 15-50, 20-50, 25-50, 130-50, 35-50, 40-50, or 45-50 mM MgCl 2 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-45, 0.1-40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-15, 0.1-10, 0.1-5, 0.1-4, 0.1-4.9, 0.1-4.8, 0.1-4.7, 0.1-4.6, 0.1-4.5, 0.1-4.4, 0.1-4.3, 0.1-4.2, 0.1-4.1, 0.1-4.0, 0.1-3.9, 0.1-3.8, 0.1-3.7, 0.1-3.6, 0.1- 3.5, 0.1-3.4, 0.1-3.3, 0.1-3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.8,
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl 2 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl 2 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl 2 .
  • the ASO is solubilized or diluted in a buffer further comprising 1-100 mM NaHCO 3 , 1-100 mM KHCO 3 , or a combination thereof.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100 mM NaHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-99, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM NaHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-100, 3-100, 4-100, 5-100, 6-100, 7- 100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60- 100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM NaHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.0-28.0, 24.0-27.9, 24.0- WSGR Docket No.47991-744.601 27.8, 24.0-27.7, 24.0-27.6, 24.0-27.5, 24.0-27.4, 24.0-27.3, 24.0-27.2, 24.0-27.1, 24.0-27.0, 24.0-26.9, 24.0-26.8, 24.0-26.7, 24.0-26.6, 24.0-26.5, 24.0-26.4, 24.0-26.3, 24.0-26.2, 24.0-26.1, 24.0-26.0, 24.0- 25.9, 4.0-25.8, 24.0-25.7, 24.0-25.6, 24.0-25.5, 24.0-25.4, 24.0-25.3, 24.0-25.2, 24.0-25.1, 24.0-25.0, 24.0-24.9, 24.0-24.8, 24.0-24.7, 24.0-24.6, 24.0-24.5, 24.0-24.4, 24.0-24.3, 24.0-24.2, or 24.0-24.1 m
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.1-28.0, 24.2-28.0, 24.3-28.0, 24.4-28.0, 24.5-28.0, 24.6-28.0, 24.7-28.0, 24.8-28.0, 24.9- 28.0, 25.0-28.0, 25.1-28.0, 25.2-28.0, 25.3-28.0, 25.4-28.0, 25.5-28.0, 25.6-28.0, 25.7-28.0, 25.8-28.0, 25.9-28.0, 26.0-28.0, 26.1-28.0, 26.2-28.0, 26.3-28.0, 26.4-28.0, 26.5-28.0, 26.6-28.0, 26.7-28.0, 26.8- 28.0, 26.9-28.0, 27.0-28.0, 27.1-28.0, 27.2-28.0, 27.3-28.0, 27.4-28.0, 27.5-28.0, 27.6-28.0, 27.7-28.0, 27.8-28.0, or 27.9-28.0 mM NaHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.0, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100 mM KHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-99, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM KHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-100, 3-100, 4-100, 5-100, 6-100, 7- 100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60- 100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM KHCO 3 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.0-28.0, 24.0-27.9, 24.0-27.8, 24.0-27.7, 24.0-27.6, 24.0-27.5, 24.0-27.4, 24.0-27.3, 24.0-27.2, 24.0-27.1, 24.0-27.0, 24.0-26.9, 24.0- 26.8, 24.0-26.7, 24.0-26.6, 24.0-26.5, 24.0-26.4, 24.0-26.3, 24.0-26.2, 24.0-26.1, 24.0-26.0, 24.0-25.9, 4.0-25.8, 24.0-25.7, 24.0-25.6, 24.0-25.5, 24.0-25.4, 24.0-25.3, 24.0-25.2, 24.0-25.1, 24.0-25.0, 24.0- 24.9, 24.0-24.8, 24.0-24.7, 24.0-24.6, 24.0-24.5, 24.0-24.4, 24.0-24.3, 24.0-24.2, or 24.0-24.1 mM KHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.1-28.0, 24.2-28.0, 24.3-28.0, 24.4-28.0, 24.5-28.0, 24.6-28.0, 24.7-28.0, 24.8-28.0, 24.9- 28.0, 25.0-28.0, 25.1-28.0, 25.2-28.0, 25.3-28.0, 25.4-28.0, 25.5-28.0, 25.6-28.0, 25.7-28.0, 25.8-28.0, 25.9-28.0, 26.0-28.0, 26.1-28.0, 26.2-28.0, 26.3-28.0, 26.4-28.0, 26.5-28.0, 26.6-28.0, 26.7-28.0, 26.8- 28.0, 26.9-28.0, 27.0-28.0, 27.1-28.0, 27.2-28.0, 27.3-28.0, 27.4-28.0, 27.5-28.0, 27.6-28.0, 27.7-28.0, WSGR Docket No.47991-744.601 27.8-28.0, or 27.9-28.0 mM KHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.0, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM KHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM KHCO3.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM KHCO3.
  • a buffer comprising 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1,
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-50 mM KH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-100, 0.01-100, 0.02-100, 0.03-100, 0.04-100, 0.05-100, 0.06-100, 0.07-100, 0.08-100, 0.09- 100, 0.1-100, 0.2-100, 0.3-100, 0.4-100, 0.5-100, 0.6-100, 0.7-100, 0.8-100, 0.9-100, 1-100, 2-100, 3- 100, 4-100, 5-100, 6-100, 7-100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100,
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-95, 0-90, 0-85, 0-80, 0-75, 0-70, 0-65, 0-60, 0-55, 0-50, 0-45, 0-40, 0-35, 0-30, 0-25, 0-20, 0-15, 0-10, 0-9, 0-8, 0-7, 0-6, 0-5, 0-4, 0-3, 0-2, 0-1, 0-0.9, 0-0.8, 0-0.7, 0-0.6, 0-0.5, 0-0.4, 0-0.3, 0-0.2, 0-0.1, 0-0.09, 0-0.08, 0-0.07, 0-0.06, 0-0.05, 0-0.04, 0-0.03, or 0-0.02 mM KH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.01-95, 0.01- 90, 0.01-85, 0.01-80, 0.01-75, 0.01-70, 0.01-65, 0.01-60, 0.01-55, 0.01-50, 0.01-45, 0.01-40, 0.01-35, 0.01-30, 0.01-25, 0.01-20, 0.01-15, 0.01-10, 0.01-9, 0.01-8, 0.01-7, 0.01-6, 0.01-5, 0.01-4, 0.01-3, 0.01-2, 0.01-1, 0.01-0.9, 0.01-0.8, 0.01-0.7, 0.01-0.6, 0.01-0.5, 0.01-0.4, 0.01-0.3, 0.01-0.2, 0.01-0.1, 0.01-0.09, 0.01-0.08, 0.01-0.07, 0.01-0.06, 0.01-0.05, 0.01-0.04, 0.01-0.03, or 0.01-0.02 mM KH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-3.0, 0-2.9, 0-2.8, 0-2.7, 0-2.6, 0-2.5, 0-2.4, 0-2.3, 0-2.2, 0-2.1, 0-2.0, 0-1.9, 0-1.8, 0-1.7, 0-1.6, 0-1.5, 0-1.4, 0-1.3, 0- 1.2, 0-1.1, 0-1.0, 0-0.9, 0-0.8, 0-0.7, 0-0.6, 0-0.5, 0-0.4, 0-0.3, or 0-0.2 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-3.0, 0.1- 2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1- 0.4, 0.1-0.3, or 0.1-0.2 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-3.0, 0.1-3.0, 0.2-3.0, 0.3-3.0, 0.4-3.0, 0.5-3.0, 0.6-3.0, 0.7-3.0, 0.8-3.0, 0.9-3.0, 1.0-3.0, 1.2-3.0, 1.3-3.0, 1.4-3.0, 1.5-3.0, 1.6-3.0, 1.7-3.0, 1.8-3.0, 1.9-3.0, 2.0-3.0, 2.1-3.0, 2.2- 3.0, 2.3-3.0, 2.4-3.0, 2.5-3.0, 2.6-3.0, 2.7-3.0, 2.8-3.0, or 2.9-3.0 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0, 0.1, 0.2, 0.3, 0.4, 0.5, WSGR Docket No.47991-744.601 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, or 3.0 mM KH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-50 mM NaH2PO4.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-50, 0-45, 0-40, 0-35, 0-30, 0-25, 0-20, 0-19, 0-18, 0-17, 0-16, 0-15, 0-14, 0-13, 0-12, 0-11, 0-10, 0-9, 0-8, 0-7, 0-6, 0-5, 0-4, 0-3, 0-2, 0-1, 0-0.9, 0-0.8, 0-0.7, 0-0.6, 0-0.5, 0-0.4, 0-0.3, or 0-0.2 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0.1-50, 0.1-45, 0.1-40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-19, 0.1-18, 0.1-17, 0.1-16, 0.1-15, 0.1-14, 0.1-13, 0.1-12, 0.1-11, 0.1-10, 0.1-9, 0.1-8, 0.1-7, 0.1-6, 0.1-5, 0.1-4, 0.1-3, 0.1-2, 0.1-1, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-50, 0.1-50, 0.2-50, 0.3-50, 0.4- 50, 0.5-50, 0.6-50, 0.7-50, 0.8-50, 0.9-50, 1-50, 2-50, 3-50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 1-50, 11- 50, 12-50, 13-50, 14-50, 15-50, 16-50, 17-50, 18-50, 19-50, 20-50, 25-50, 30-50, 35-50, 40-50, or 45-50 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0-20, 0.1-20, 0.2-20, 0.3-20, 0.4-20, 0.5-20, 0.6-20, 0.7-20, 0.8-20, 0.9-20, 1-20, 2-20, 3-20, 4-20, 5-20, 6-20, 7-20, 8-20, 9-20, 10-20, 11-20, 12-20, 13-20, 14-20, 15-20, 16-20, 17-20, 18-20, or 19- 20 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM NaH 2 PO 4 .
  • the ASO as described herein is solubilized or diluted in a buffer comprising 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM NaH 2 PO 4 .
  • the ASO is solubilized or diluted in a buffer further comprising carbohydrates.
  • the carbohydrates comprise D-glucose.
  • the ASO is solubilized or diluted in a buffer further comprising 1-100 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 1-100, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-29, 1-28, 1-27, 1-26, 1-25, 1-24, 1-23, 1-22, 1-21, 1-20, 1-19, 1-18, 1-17, 1-16, 1-15, 1-14, 1-13, 1-12, 1-11, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM D-glucose.
  • the ASO as described WSGR Docket No.47991-744.601 herein is solubilized or diluted in a buffer comprising 2-100, 3-100, 4-100, 5-100, 6-100, 7-100, 8-100, 9- 100, 10-100, 11-100, 12-100, 13-100, 14-100, 15-100, 16-100, 17-100, 18-100, 19-100, 20-100, 21-100, 22-100, 23-100, 24-100, 25-100, 26-100, 29-100, 28-100, 29-100, 30-100, 35-100, 40-100, 45-100, 50- 100, 55-100, 60-100, 65-100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 2-30, 3- 30, 4-30, 5-30, 6-30, 7-30, 8-30, 9-30, 10-30, 11-30, 12-30, 13-30, 14-30, 15-30, 16-30, 17-30, 18-30, 19- 30, 20-30, 21-30, 22-30, 23-30, 24-30, 25-30, 26-30, 27-30, 28-30, or 29-30 mM D-glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM D- glucose.
  • the ASO as described herein is solubilized or diluted in a buffer comprising at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM D-glucose. In some embodiments, the ASO as described herein is solubilized or diluted in a buffer comprising 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 mM D-glucose.
  • the ASO is solubilized or diluted in a buffer comprising 25-250 mM NaCl, 0.1-20 mM KCl, 0.1-50 mM Na 2 HPO 4 , 0.1-50 mM NaH 2 PO 4 , 0.1-50 mM CaCl 2 , and 0.1-50 mM MgCl 2 .
  • the ASO is solubilized or diluted in a buffer comprising 150 mM NaCl, 3.0 mM KCl, 0.7 mM Na 2 HPO 4 , 0.3 mM NaH 2 PO 4 , 0.79 mM MgCl 2 , and 1.4 mM CaCl 2 .
  • the ASO is solubilized or diluted in a buffer further comprising an antioxidant.
  • the antioxidant is t-butylhydroxyquinoline (TBHQ), butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), vitamin E, or any combination thereof.
  • the ASO is solubilized or diluted in a buffer further comprising an antioxidant, wherein the antioxidant is ascorbic acid (vitamin C), glutathione, lipoic acid, uric acid, carotenes, ⁇ -tocopherol (vitamin E), ubiquinol (coenzyme Q), or any combination thereof.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-250 mM NaCl, 0.1-20 mM KCl, 0-50 mM KH 2 PO4, 1-100 mM NaHCO 3 , 0-50 mM NaH2PO4, 1-100 mM D-glucose, 0.1-50 mM CaCl 2 , 0.1-50 mM MgCl 2 , or any combination thereof.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 25-250 mM NaCl, 0.1-20 mM KCl, 0-50 mM KH2PO4, 1-100 mM NaHCO3, 0-50 mM NaH2PO4, 1-100 mM D-glucose, 0.1-50 mM CaCl2, and 0.1-50 mM MgCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 127 mM NaCl, 1.0 mM KCl, 1.2 mM KH2PO4, 26 mM NaHCO3, 10 mM D-glucose, 2.4 mM CaCl2, and 1.3 mM MgCl2.
  • the ASO as described herein is solubilized or diluted in a buffer comprising 119 mM NaCl, 26.2 mM NaHCO3, 2.5 mM KCl, 1 mM NaH2PO4, 1.3 mM MgCl2, 10 mM glucose, and 2.5 mM CaCl2.
  • the pharmaceutical formulation does not comprise a preservative. In some embodiments, the pharmaceutical formulation comprises a preservative. [0417] In some embodiments, the ASO as described herein is solubilized or diluted to a concentration of from 5-250 mg/mL in the diluent.
  • the ASO as described herein is solubilized or diluted to a concentration of from 5-250, 5-247.5, 5-245, 5-242.5, 5-240, 5-237.5, 5-235, 5-232.5, 5-230, 5-227.5, 5-225, 5-225.5, 5- 220, 5-217.5, 5-215, 5-212.5, 5-210, 5-205.5, 5-205, 5-202.5, 5-200, 5-197.5, 5-195, 5-192.5, 5-190, 5- 187.5, 5-185, 5-182.5, 5-180, 5-177.5, 5-175, 5-172.5, 5-170, 5-167.5, 5-165, 5-162.5, 5-160, 5-157.5, 5- 155, 5-152.5, 5-150, 5-147.5, 5-145, 5-142.5, 5-140, 5-137.5, 5-135, 5-132.5, 5-130, 5-127.5, 5-125, 5- 122.5, 5-120, 5-117.5, 5-115, 5-112.5, 5-110, 5-107.5, 5-105, 5-102.5, 5-100, 5-97.5, 5-95,
  • the ASO as described herein is solubilized or diluted to a concentration of from 10-250, 15-250, 20-250, 25-250, 30-250, 35-250, 40-250, 45-250, 50- 250, 55-250, 60-250, 65-250, 70-250, 75-250, 80-250, 85-250, 90-250, 95-250, 100-250, 105-250, 110- 250, 115-250, 120-250, 125-250, 130-250, 135-250, 140-250, 145-250, 150-250, 155-250, 160-250, 165- 250, 170-250, 175-250, 180-250, 185-250, 190-250, or 195-250, 200-250, 205-250, 210-250, 215-250, 220-250, 225-250, 230-250, 235-250, 240-250, or 245-250 mg/mL in the diluent.
  • the ASO as described herein is solubilized or diluted to a concentration of from at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109,
  • the ASO as described herein is solubilized or diluted to a concentration of from at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109,
  • the ASO as described herein is solubilized or diluted to a concentration of from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110,
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of the ASO is present in the pharmaceutical composition at a concentration of from 0.1 mg/mL to 250 mg/mL. In some embodiments, the ASO as described herein is present in the pharmaceutical composition at a concentration of about 0.1 mg/mL, 0.5 mg/mL, 1 mg/mL, 2 mg/mL, 2.5 mg/mL, 3 mg/mL, 4 mg/mL, 5 mg/mL, 6 mg/mL, 7 mg/mL, 8 mg/mL, 9 mg/mL, 10 mg/mL, 11 mg/mL, 12 mg/mL, 13 mg/mL, 14 mg/mL, 15 mg/mL, 16 mg/mL, 17 mg/mL, 18 mg/mL, 19 mg/mL, or 20 mg/mL.
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of about 30 mg/mL, 40 mg/mL, 50 mg/mL, 60 mg/mL, 70 mg/mL, 80 mg/mL, 90 mg/mL, 100 mg/mL, 110 mg/mL, 120 mg/mL, 130 mg/mL, 140 mg/mL, 150 mg/mL, 160 mg/mL, 170 mg/mL, 180 mg/mL, 190 mg/mL, or 200 mg/mL.
  • the ASO is present in the pharmaceutical composition at a concentration of about 22.5 mg/mL, 25 mg/mL, 27.5 mg/mL, 30 mg/mL, 32.5 mg/mL, 35 mg/mL, 37.5 mg/mL, 40 mg/mL, 42.5 mg/mL, 45 mg/mL, 47.5 mg/mL, 50 mg/mL, 52.5 mg/mL, 55 mg/mL, 57.5 mg/mL, 60 mg/mL, 62.5 mg/mL, 65 mg/mL, 67.5 mg/mL, 70 mg/mL, 72.5 mg/mL, 75 mg/mL, 77.5 mg/mL, 80 mg/mL, 82.5 mg/mL, 85 mg/mL, 87.5 mg/mL, 90 mg/mL, 92.5 mg/mL, 95 mg/mL, 97.5 mg/mL, 100 mg/mL, 102.5 mg/mL, 105 mg/mL, 107.5 mg/mL, 110 mg/mL,
  • WSGR Docket No.47991-744.601 the ASO as described herein is solubilized or diluted to a concentration of 11 mg/mL, 22 mg/mL, 33 mg/mL, 44 mg/mL, 55 mg/mL, 66 mg/mL, 77 mg/mL, 88 mg/mL, 99 mg/mL, or 100 mg/mL in the diluent.
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 0.1 mg/mL to 250 mg/mL, from 0.2 mg/mL to 250 mg/mL, from 0.3 mg/mL to 250 mg/mL, from 0.4 mg/mL to 250 mg/mL, from 0.5 mg/mL to 250 mg/mL, from 0.6 mg/mL to 250 mg/mL, from 0.7 mg/mL to 250 mg/mL, from 0.8 mg/mL to 250 mg/mL, from 0.9 mg/mL to 250 mg/mL, from 1.0 mg/mL to 250 mg/mL, from 1.1 mg/mL to 250 mg/mL, from 1.2 mg/mL to 250 mg/mL, from 1.3 mg/mL to 250 mg/mL, from 1.4 mg/mL to 250 mg/mL, from 1.5 mg/mL to 250 mg/mL, from 1.6 mg/mL to 250 mg/mL, from 1.7 mg/
  • the ASO as described herein is present in the pharmaceutical composition at a concentration of from 0.1 mg/mL to 250 mg/mL, from 0.1 mg/mL to 245 mg/mL, from 0.1 mg/mL to 240 mg/mL, from 0.1 mg/mL to 235 mg/mL, from 0.1 mg/mL to 230 mg/mL, from 0.1 mg/mL to 225 WSGR Docket No.47991-744.601 mg/mL, from 0.1 mg/mL to 220 mg/mL, from 0.1 mg/mL to 215 mg/mL, from 0.1 mg/mL to 210 mg/mL, from 0.1 mg/mL to 205 mg/mL, from 0.1 mg/mL to 200 mg/mL, from 0.1 mg/mL to 195 mg/mL, from 0.1 mg/mL to 190 mg/mL, from 0.1 mg/mL to 185 mg/mL, from 0.1 mg/
  • the ASO as described herein is solubilized or diluted to a concentration of at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57,
  • the ASO as described herein is solubilized or diluted to a concentration of at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58,
  • the ASO as described herein is solubilized or diluted to a concentration of 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59,
  • the ASO as described herein is formulated in a dose volume adjusted depending on the subject’s age, sex, and/or body mass index (BMI). In some embodiments, the ASO as described herein is formulated in a dose volume of 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mL.
  • the ASO as described herein is formulated in a dose volume of 0.5, 0.75, 1.0, 1.25, 1.5, 1.75, 2, 2.25, 2.5.2.75, 3, 3.25, 3.5, 3.75, 4, 4.25, 4.5, 4.75, 5, 5.25, 5.5, 5.75, 6, 6.25, 6.5, 6.75, 7, 7.25, 7.5, 7.75, 8, 8.25, 8.5, 8.75, 9, 9.25, 9.5, or 10 mL.
  • the ASO as described herein is formulated in a dose volume of less than 20 mL, less than 19 mL, less than 18 mL, less than 17 mL, less than 16 mL, less than 15 mL, less than 14 mL, less than 13 mL, less than 12 WSGR Docket No.47991-744.601 mL, less than 11 mL, less than 10 mL, less than 9 mL, less than 8 mL, less than 7 mL, less than 6 mL, less than 5 mL, less than 4 mL, less than 3 mL, less than 2 mL, less than 1 mL.
  • the ASO as described herein is formulated in a dose volume of more than 1 mL, more than 2 mL, more than 3 mL, more than 4 mL, more than 5 mL, more than 6 mL, more than 7 mL, more than 8 mL, more than 9 mL, more than 10 mL, more than 11 mL, more than 12 mL, more than 13 mL, more than 14 mL, more than 15 mL, more than 16 mL, more than 17 mL, more than 18 mL, more than 19 mL, more than 20 mL.
  • the ASO as described herein is formulated in a dose volume of at least 0.5 mL, at least 1 mL, at least 2 mL, at least 3 mL, at least 4 mL, at least 5 mL, at least 6 mL, at least 7 mL, at least 8 mL, at least 9 mL, at least 10 mL, at least 11 mL, at least 12 mL, at least 13 mL, at least 14 mL, at least 15 mL, at least 16 mL, at least 17 mL, at least 18 mL, at least 19 mL, at least 20 mL.
  • the ASO as described herein is formulated in a final dose volume adjusted depending on the subject’s age, sex, and/or body mass index (BMI). In some embodiments, the ASO as described herein is formulated in a final dose volume of 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mL.
  • the ASO as described herein is formulated in a final dose volume of 0.5, 0.75, 1.0, 1.25, 1.5, 1.75, 2, 2.25, 2.5.2.75, 3, 3.25, 3.5, 3.75, 4, 4.25, 4.5, 4.75, 5, 5.25, 5.5, 5.75, 6, 6.25, 6.5, 6.75, 7, 7.25, 7.5, 7.75, 8, 8.25, 8.5, 8.75, 9, 9.25, 9.5, or 10 mL.
  • the ASO as described herein is formulated in a final dose volume of less than 20 mL, less than 19 mL, less than 18 mL, less than 17 mL, less than 16 mL, less than 15 mL, less than 14 mL, less than 13 mL, less than 12 mL, less than 11 mL, less than 10 mL, less than 9 mL, less than 8 mL, less than 7 mL, less than 6 mL, less than 5 mL, less than 4 mL, less than 3 mL, less than 2 mL, less than 1 mL.
  • the ASO as described herein is formulated in a final dose volume of more than 1 mL, more than 2 mL, more than 3 mL, more than 4 mL, more than 5 mL, more than 6 mL, more than 7 mL, more than 8 mL, more than 9 mL, more than 10 mL, more than 11 mL, more than 12 mL, more than 13 mL, more than 14 mL, more than 15 mL, more than 16 mL, more than 17 mL, more than 18 mL, more than 19 mL, more than 20 mL.
  • the ASO as described herein is formulated in a final dose volume of at least 0.5 mL, at least 1 mL, at least 2 mL, at least 3 mL, at least 4 mL, at least 5 mL, at least 6 mL, at least 7 mL, at least 8 mL, at least 9 mL, at least 10 mL, at least 11 mL, at least 12 mL, at least 13 mL, at least 14 mL, at least 15 mL, at least 16 mL, at least 17 mL, at least 18 mL, at least 19 mL, at least 20 mL.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: i) an active pharmaceutical ingredient, and ii) a pharmaceutically acceptable diluent; wherein the liquid composition is not buffered by a buffering agent; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, and/or (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising:(i) an active pharmaceutical ingredient, and (ii) a pharmaceutically acceptable diluent; wherein the liquid composition lacks Na2HPO4 and/or WSGR Docket No.47991-744.601 NaH2PO4; and wherein: (a) the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion, and/or (b) the pharmaceutical formulation is formulated or suitable for administration into an intrathecal space, cerebrospinal fluid, or a brain of a human subject.
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • a pharmaceutical formulation wherein the pharmaceutical formulation is a liquid composition comprising: (i) an active pharmaceutical ingredient, and (ii) a pharmaceutically acceptable diluent consisting of: a) NaCl, b) KCl, c) MgCl2 or MgCl26H2O, and d) CaCl2 or CaCl22H2O.
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • the pharmaceutically acceptable diluent provided herein is suitable for preparation of any pharmaceutical formulation intrathecal administration.
  • any active pharmaceutical ingredient suitable for intrathecal administration may be formulated using the pharmaceutically acceptable diluent provided herein.
  • the active pharmaceutical ingredient can be a small molecule or a biologic, such as an organic chemical compound, an antisense oligonucleotide, a DNA, an antibody or any other protein or polypeptide, a living organism such as bacterium or fungi, viral vector, or viral-like particle, or any combination thereof.
  • the pharmaceutical formulation does not comprise a preservative.
  • the pharmaceutical formulation comprises a preservative.
  • a pharmaceutical composition used in the therapeutic methods of the invention is formulated to be compatible with its intended route of administration.
  • the pharmaceutical formulation is suitable for an intracerebroventricular or intrathecal injection.
  • the pharmaceutical formulation is suitable for oral, rectal, intranasal, intradermal, subcutaneous, intrathecal, intracerebroventricular, intraperitoneal, intramuscular, intravitreal, intravenous, intracranial, intrabuccal, or sublingual administration.
  • the pharmaceutical formulation is suitable for intradermal, subcutaneous, intrathecal, intranasal, intracranial, intracerebroventricular, intraperitoneal, intramuscular, intravitreal, or intravenous injection.
  • the pharmaceutical formulation is packaged in a single-use vial. In some embodiments, the pharmaceutical formulation is packaged in a multiple use vial.
  • Pharmaceutical formulations comprising the agent, e.g., antisense oligonucleotide or antisense oligomer, of the described compositions and for use in any of the described methods can be prepared according to conventional techniques well-known in the pharmaceutical industry and described in the published literature.
  • a pharmaceutical formulation for treating a subject comprises an effective amount of any antisense oligomer as described herein, or a pharmaceutically acceptable salt, solvate, hydrate or ester thereof.
  • the pharmaceutical composition described herein further comprises a pharmaceutically acceptable excipient, carrier, or diluent.
  • a pharmaceutically acceptable excipient such as neutral buffered saline, phosphate buffered saline and the like; carbohydrates such as glucose, mannose, sucrose or dextrans, mannitol; proteins; polypeptides or amino acids such as glycine; antioxidants; chelating agents such as EDTA or glutathione; adjuvants (e.g., aluminum hydroxide); and preservatives.
  • a pharmaceutical composition used in the therapeutic methods of the invention is formulated to be compatible with its intended route of administration.
  • compositions and “pharmaceutical formulation” (or “formulation”) are used interchangeably and denote a mixture or solution comprising a therapeutically effective amount of an active pharmaceutical ingredient together with one or more pharmaceutically acceptable excipients to be administered to a subject, e.g., a human in need thereof.
  • compositions are formulated into any of many possible dosage forms such as, but not limited to, solutions, liquids, tablets, capsules, gel capsules, liquid syrups, soft gels, suppositories, and enemas.
  • the compositions are formulated as suspensions in aqueous, non-aqueous or mixed media.
  • Aqueous suspensions may further contain substances that increase the viscosity of the suspension including, for example, sodium carboxymethylcellulose, sorbitol and/or dextran.
  • the suspension may also contain stabilizers.
  • a pharmaceutical formulation or composition of the present invention includes, but is not limited to, a solution, emulsion, microemulsion, foam or liposome-containing formulation (e.g., cationic or noncationic liposomes).
  • the pharmaceutical composition or formulation described herein may comprise one or more penetration enhancers, carriers, excipients or other active or inactive ingredients as appropriate and well- known to those of skill in the art or described in the published literature.
  • liposomes also include sterically stabilized liposomes, e.g., liposomes comprising one or more specialized lipids. These specialized lipids result in liposomes with enhanced circulation lifetimes.
  • a sterically stabilized liposome comprises one or more glycolipids or is derivatized with one or more hydrophilic polymers, such as a polyethylene glycol (PEG) moiety.
  • PEG polyethylene glycol
  • a surfactant is included in the pharmaceutical formulation or compositions. The use of surfactants in drug products, formulations and emulsions is well-known in the art.
  • the present invention employs a penetration enhancer to effect the efficient delivery of the antisense oligonucleotide or antisense oligomer, e.g., to aid diffusion across cell membranes and/or enhance the permeability of a lipophilic drug.
  • the penetration enhancers are a surfactant, fatty acid, bile salt, chelating agent, or non-chelating nonsurfactant.
  • the pharmaceutical formulation comprises multiple antisense oligonucleotides or antisense oligomers.
  • the antisense oligonucleotide or antisense oligomer is administered in combination with another drug or therapeutic agent.
  • compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion.
  • Solutions or suspensions used for parenteral, intranasal, intradermal, or subcutaneous application can include the following components: a sterile diluent such as water for WSGR Docket No.47991-744.601 injection, saline solution, fixed oils, polyethylene glycols, glycerin, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose.
  • a sterile diluent such as water for WSGR
  • the vehicle may contain water, synthetic or vegetable oil, and/or organic co-solvents.
  • the parenteral formulation would be reconstituted or diluted prior to administration. pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide.
  • Depot formulations providing controlled or sustained release of an invention composition, may include injectable suspensions of nano/micro particles or nano/micro or non-micronized crystals.
  • suitable carriers include physiological saline, bacteriostatic water, Cremophor ELTM (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS).
  • the carrier can be a solvent or dispersion medium containing, for example, water, ethanol, poly(ol) (for example, glycerol, propylene glycol, and liquid polyetheylene glycol, and the like), and suitable mixtures thereof.
  • the proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants.
  • Sterile injectable solutions can be prepared by incorporating the composition in the required amount in an appropriate solvent with one or a com-bination of ingredients enumerated above, as required, followed by filtered sterilization. Prevention of the action of micro-organisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like.
  • dispersions are prepared by incorporating the active composition into a sterile vehicle which contains a basic dispersion medium and the required other ingredients from those enumerated above.
  • the preferred methods of preparation are vacuum drying and freeze-drying which yields a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof.
  • isotonic agents for example, sugars, polyalcohols such as mannitol, sorbitol, and sodium chloride in the composition.
  • Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate and gelatin.
  • the parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic.
  • compositions can be formulated in liquid or solid dosage forms and as instant or controlled/sustained release formulations.
  • Suitable dosage forms for oral ingestion by a subject include powders, tablets, pills, granules, dragees, hard- and soft-shell capsules, liquids, gels, syrups, slurries, suspensions, emulsions, and the like.
  • Oral compositions generally include an inert WSGR Docket No.47991-744.601 diluent or an edible carrier. They can be enclosed in gelatin capsules or compressed into tablets.
  • the active agent can be incorporated with excipients and used in the form of tablets, troches, or capsules.
  • Oral compositions can also be prepared using a fluid carrier for use as a mouthwash, wherein the agent in the fluid carrier is applied orally and swished and expectorated or swallowed.
  • Pharmaceutically compatible binding agents, and/or adjuvant materials can be included as part of the composition.
  • the tablets, granules, pills, capsules, troches and the like can contain any of the following ingredients, or compounds of a similar nature: a binder such as micro- crystalline cellulose, gum tragacanth or gelatin; an excipient such as starch or lactose; dissolution retardant; anti-adherents; cationic exchange resin; wetting agents; antioxidants; preservatives; a disintegrating agent such as alginic acid, Primogel, or corn starch; a lubricant such as magnesium stearate or Sterotes; a glidant such as colloidal silicon dioxide; a preservative; a colorant; a sweetening agent such as sugars such as dextrose, sucrose or saccharin; or a flavoring agent such as peppermint, methyl salicylate, or orange flavoring, each of these being synthetic and/or natural.
  • a binder such as micro- crystalline cellulose, gum tragacanth or gelatin
  • an excipient
  • compositions are delivered in the form of an aerosol spray from pressured container or dispenser which contains a suitable propellant, e.g., a gas such as carbon dioxide, or a nebulizer.
  • a suitable propellant e.g., a gas such as carbon dioxide, or a nebulizer.
  • Systemic administration can also be by transmucosal or trans-dermal means.
  • penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives.
  • Transmucosal administration can be accomplished through the use of nasal sprays or suppositories.
  • the active agents are formulated into ointments, salves, gels, or creams, emulsion, a solution, a suspension, or a foam, as generally known in the art.
  • the penetration of the drug into the skin and underlying tissues can be regulated, for example, using penetration enhancers; the appropriate choice and combination of lipophilic, hydrophilic, and amphiphilic excipients, including water, organic solvents, waxes, oils, synthetic and natural polymers, surfactants, emulsifiers; by pH adjustments; use of complexing agents and other techniques, such as iontophoresis, may be used to regulate skin penetration of the active ingredient.
  • the compositions may also be formulated in rectal compositions, such as suppositories (e.g., with conventional suppository bases such as cocoa butter and other glycerides) or retention enemas.
  • Examples of pharmaceutically or physiologically acceptable carriers, diluents or excipients include, but are not limited to, antifoaming agents, antioxidants, binders, carriers or carrier materials, dispersing agents, viscosity modulating agents, diluents, filling agents, lubricants, glidants, plasticizers, solubilizers, stabilizers, suspending agents, surfactants, viscosity enhancing agents, and wetting agents.
  • the separate components of the compositions of the invention may be preblended or each component may be added separately to the same environment according to a predetermined dosage for the purpose of achieving the desired concentration level of the treatment components and so long as the components eventually come into intimate admixture with each other.
  • the invention may be administered or delivered on a continuous or intermittent basis.
  • WSGR Docket No.47991-744.601 [0450] It is especially advantageous to formulate oral or parenteral compositions in dosage unit form for ease of administration and uniformity of dosage.
  • Dosage unit form as used herein refers to physically discrete units suited as unitary dosages for the subject to be treated; each unit containing a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier.
  • the specification for the dosage unit forms of the invention is dictated by and directly dependent on the unique characteristics of the compositions and the particular therapeutic effect to be achieved, and the limitations inherent in the art of compounding such an agent for the treatment of subjects.
  • the ASOs disclosed in the present disclosure can be used in combination with one or more additional therapeutic agents.
  • the one or more additional therapeutic agents can comprise a small molecule.
  • the one or more additional therapeutic agents can comprise a small molecule described in WO2016128343A1, WO2017053982A1, WO2016196386A1, WO201428459A1, WO201524876A2, WO2013119916A2, and WO2014209841A2, which are incorporated by reference herein in their entirety.
  • the one or more additional therapeutic agents comprise an ASO that can be used to correct intron retention.
  • the one or more other agents are selected from the ASOs listed in Table 8A or Table 8B.
  • Treatment of Subjects Any of the compositions provided herein may be administered to an individual. “Individual” may be used interchangeably with “subject” or “patient.”
  • An individual may be a mammal, for example a human or animal such as a non-human primate, a rodent, a rabbit, a rat, a mouse, a horse, a donkey, a goat, a cat, a dog, a cow, a pig, or a sheep. In embodiments, the individual is a human.
  • the individual is a fetus, an embryo, or a child. In other embodiments, the individual may be another eukaryotic organism, such as a plant.
  • the compositions provided herein are administered to a cell ex vivo. [0454] In some embodiments, the compositions provided herein are administered to an individual as a method of treating a disease or disorder. In some embodiments, the individual has a genetic disease, such as any of the diseases described herein. In some embodiments, the individual is at risk of having a disease, such as any of the diseases described herein. In some embodiments, the individual is at increased risk of having a disease or disorder caused by insufficient amount of a protein or insufficient activity of a protein.
  • an individual is “at an increased risk” of having a disease or disorder caused insufficient amount of a protein or insufficient activity of a protein
  • the method involves preventative or prophylactic treatment.
  • an individual may be at an increased risk of having such a disease or disorder because of family history of the disease.
  • individuals at an increased risk of having such a disease or disorder benefit from prophylactic treatment (e.g., by preventing or delaying the onset or WSGR Docket No.47991-744.601 progression of the disease or disorder).
  • a fetus is treated in utero, e.g., by administering the ASO composition to the fetus directly or indirectly (e.g., via the mother).
  • Suitable routes for administration of ASOs of the present disclosure may vary depending on cell type to which delivery of the ASOs is desired. Multiple tissues and organs are affected by Dravet syndrome; Epilepsy, generalized, with febrile seizures plus, type 2; Febrile seizures, familial, 3A; Migraine, familial hemiplegic, 3; Autism; Epileptic encephalopathy, early infantile, 13; Sick sinus syndrome 1; Alzheimer’s disease or SUDEP, with the brain being the most significantly affected tissue.
  • the ASOs of the present disclosure may be administered to patients parenterally, for example, by intrathecal injection, intracerebroventricular injection, intraperitoneal injection, intramuscular injection, subcutaneous injection, intravitreal injection, or intravenous injection.
  • the method comprising administering to the subject the pharmaceutical composition comprising the ASO as described herein.
  • the subject is a human subject.
  • Therapeutic Target Population the human subject is at most 18 years old at a first dose of the pharmaceutical composition.
  • the human subject is from 1 to 18, from 2 to 18, from 3 to 18, from 4 to 18, from 5 to 18, from 6 to 18, from 7 to 18, from 8 to 18, from 9 to 18, from 10 to 18, from 11 to 18, from 12 to 18, from 13 to 18, from 14 to 18, from 15 to 18, from 16 to 18, or from 17 to 18 years old.
  • the human subject is a human from 1 to 17, from 1 to 16, from 1 to 15, from 1 to 14, from 1 to 13, from 1 to 12, from 1 to 11, from 1 to 10, from 1 to 9, from 1 to 8, from 1 to 7, from 1 to 6, from 1 to 5, from 1 to 4, from 1 to 3, or from 1 to 2 years old. In some embodiments, the human subject is less than a year old or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 years old.
  • the human subject is at most 35 years, 30 years, 29 years, 28 years, 27 years, 26 years, 25 years, 24 years, 23 years, 22 years, 21 years, 20 years, 19 years, 18 years, 17 years, 16 years, 15 years, 14 years, 13 years, 12 years, 11 years, 10 years, 9 years, 8 years, 7 years, 6 years, 5 years, 4 years, 3 years, 2 years, or 1 year old.
  • the human subject is less than a year old or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, or 35 years old.
  • the human subject is from 1 to 35, from 2 to 35, from 3 to 35, from 4 to 35, from 5 to 35, from 6 to 35, from 7 to 35, from 8 to 35, from 9 to 35, from 10 to 35, from 11 to 35, from 12 to 35, from 13 to 35, from 14 to 35, from 15 to 35, from 16 to 35, 17 to 35, from 18 to 35, from 19 to 35, from 20 to 35, from 21 to 35, from 22 to 35, from 23 to 35, from 24 to 35, from 25 to 35, from 26 to 35, from 27 to 35, from 28 to 35, from 29 to 35, from 30 to 35, from 31 to 35, from 32 to 35, from 33 to 35, or from 34 to 35 years old.
  • the human subject is a human from 1 to 35, from 1 to 34, from 1 to 33, from 1 to 32, from 1 to 31, from 1 to 30, from 1 to 29, from 1 to 28, from 1 to 27, from 1 to 26, from 1 to 25, from 1 to 24, from 1 to 23, from 1 to 22, from 1 to 21, from 1 to 20, from 1 to 19, from 1 to 18, from 1 to 17, from 1 to 16, from 1 to 15, from 1 to 14, from 1 to 13, from 1 to 12, from 1 to 11, from 1 to 10, from 1 to 9, from 1 to 8, from 1 to 7, from 1 to 6, from 1 to 5, from 1 to 4, from 1 to 3, or from 1 to 2 years old.
  • the human subject is a human from 2 to 35, from 2 to 34, from 2 to 33, from 2 to 32, from 2 to 31, from 2 to 30, from 2 to 29, from 2 to 28, from 2 to 27, from 2 to 26, from 2 to 25, from 2 to 24, from 2 to 23, from 2 to 22, from 2 to 21, from 2 to 20, from 2 to 19, from 2 to 18, from 2 to 17, from 2 to 16, from 2 to 15, from 2 to 14, from 2 to 13, from 2 to 12, from 2 to 11, from 2 to 10, from 2 to 9, from 2 to 8, from 2 to 7, from 2 to 6, from 2 to 5, from 2 to 4, or from 2 to 3 years old.
  • the human subject is a human from 3 to 35, from 3 to 34, from 3 to 33, from 3 to 32, from 3 to 31, from 3 to 30, from 3 to 29, from 3 to 28, from 3 to 27, from 3 to 26, from 3 to 25, from 3 to 24, from 3 to 23, from 3 to 22, from 3 to 21, from 3 to 20, from 3 to 19, from 3 to 18, from 3 to 17, from 3 to 16, from 3 to 15, from 3 to 14, from 3 to 13, from 3 to 12, from 3 to 11, from 3 to 10, from 3 to 9, from 3 to 8, from 3 to 7, from 3 to 6, from 3 to 5, or from 3 to 4 years old.
  • the human subject is a human from 4 to 35, from 4 to 34, from 4 to 33, from 4 to 32, from 4 to 31, from 4 to 30, from 4 to 29, from 4 to 28, from 4 to 27, from 4 to 26, from 4 to 25, from 4 to 24, from 4 to 23, from 4 to 22, from 4 to 21, from 4 to 20, from 4 to 19, from 4 to 18, from 4 to 17, from 4 to 16, from 4 to 15, from 4 to 14, from 4 to 13, from 4 to 12, from 4 to 11, from 4 to 10, from 4 to 9, from 4 to 8, from 4 to 7, from 4 to 6, or from 4 to 5 years old.
  • the human subject is a human from 5 to 35, from 5 to 34, from 5 to 33, from 5 to 32, from 5 to 31, from 5 to 30, from 5 to 29, from 5 to 28, from 5 to 27, from 5 to 26, from 5 to 25, from 5 to 24, from 5 to 23, from 5 to 22, from 5 to 21, from 5 to 20, from 5 to 19, from 5 to 18, from 5 to 17, from 5 to 16, from 5 to 15, from 5 to 14, from 5 to 13, from 5 to 12, from 5 to 11, from 5 to 10, from 5 to 9, from 5 to 8, from 5 to 7, or from 5 to 6 years old.
  • the human subject is a human from 6 to 35, from 6 to 34, from 6 to 33, from 6 to 32, from 6 to 31, from 6 to 30, from 6 to 29, from 6 to 28, from 6 to 27, from 6 to 26, from 6 to 25, from 6 to 24, from 6 to 23, from 6 to 22, from 6 to 21, from 6 to 20, from 6 to 19, from 6 to 18, from 6 to 17, from 6 to 16, from 6 to 15, from 6 to 14, from 6 to 13, from 6 to 12, from 6 to 11, from 6 to 10, from 6 to 9, from 6 to 8, or from 6 to 7 years old.
  • the human subject is a human from 7 to 35, from 7 to 34, from 7 to 33, from 7 to 32, from 7 to 31, from 7 to 30, from 7 to 29, from 7 to 28, from 7 to 27, from 7 to 26, from 7 to 25, from 7 to 24, from 7 to 23, from 7 to 22, from 7 to 21, from 7 to 20, from 7 to 19, from 7 to 18, from 7 to 17, from 7 to 16, from 7 to 15, from 7 to 14, from 7 to 13, from 7 to 12, from 7 to 11, from 7 to 10, from 7 to 9, from 7 to 8 years old.
  • the human subject is a human from 8 to 35, from 8 to 34, from 8 to 33, from 8 to 32, from 8 to 31, from 8 to 30, from 8 to 29, from 8 to 28, from 8 to 27, from 8 to 26, from 8 to WSGR Docket No.47991-744.601 25, from 8 to 24, from 8 to 23, from 8 to 22, from 8 to 21, from 8 to 20, from 8 to 19, from 8 to 18, from 8 to 17, from 8 to 16, from 8 to 15, from 8 to 14, from 8 to 13, from 8 to 12, from 8 to 11, from 8 to 10, or from 8 to 9 years old.
  • Therapeutic Dose [0469]
  • the method comprises administering the ASO at a first dose of from about 0.5 milligrams to about 500 milligrams.
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising an ASO as described herein at a first dose of about 0.1, 0.5, 1, 2.5, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, or 200 mg.
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of from about 0.1 to about 1000 mg, from about 0.2 to about 1000 mg, from about 0.3 to about 1000 mg, from about 0.4 to about 1000 mg, from about 0.5 to about 1000 mg, from about 0.6 to about 1000 mg, from about 0.7 to about 1000 mg, from about 0.8 to about 1000 mg, from about 0.9 to about 1000 mg, 1 to about 1000 mg, from about 2 to about 1000 mg, from about 3 to about 1000 mg, from about 4 to about 1000 mg, from about 5 to about 1000 mg, from about 6 to about 1000 mg, from about 7 to about 1000 mg, from about 8 to about 1000 mg, from about 9 to about 1000 mg, from about 10 to about 1000 mg, from about 15 to about 1000 mg, from about 20 to about 1000 mg, from about 25 to about 1000 mg, from about 30 to about 1000 mg, from about 35 to about 1000 mg, from about 40 to about 1000 mg, from about 45 to about 1000 mg
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of from 0.1 to 1000 mg, from 0.2 to 1000 mg, from 0.3 to 1000 mg, from 0.4 to 1000 mg, from 0.5 to 1000 mg, from 0.6 to 1000 mg, from 0.7 to 1000 mg, from 0.8 to 1000 mg, from 0.9 to 1000 mg, 1 to 1000 mg, from 2 to 1000 mg, from 3 to 1000 mg, from 4 to 1000 mg, from 5 to 1000 mg, from 6 to 1000 mg, from 7 to 1000 mg, from 8 to 1000 mg, from 9 to 1000 mg, from 10 to 1000 mg, from 15 to 1000 mg, from 20 to 1000 mg, from 25 to 1000 mg, from 30 to 1000 mg, from 35 to 1000 mg, from 40 to 1000 mg, from 45 to 1000 WSGR Docket No.47991-744.601 mg, from 50 to 1000 mg, from 55 to 1000 mg, from 60 to 1000 mg, from 65 to 1000 mg, from 70 to 1000 mg, from 75 to 1000 mg
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of from about 0.1 to about 950 mg, from about 0.1 to about 900 mg, from about 0.1 to about 850 mg, from about 0.1 to about 800 mg, from about 0.1 to about 750 mg, from about 0.1 to about 700 mg, from about 0.1 to about 650 mg, from about 0.1 to about 600 mg, from about 0.1 to about 550 mg, from about 0.1 to about 500 mg, from about 0.1 to about 450 mg, from about 0.1 to about 400 mg, from about 0.1 to about 350 mg, from about 0.1 to about 300 mg, from about 0.1 to about 250 mg, from about 0.1 to about 200 mg, from about 0.1 to about 150 mg, from about 0.1 to about 100 mg, from about 0.1 to about 95 mg, from about 0.1 to about 90 mg, from about 0.1 to about 85 mg, from about 0.1 to about 80 mg, from about 0.1 to about 75 mg,
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of from 0.1 to 950 mg, from 0.1 to 900 mg, from 0.1 to 850 mg, from 0.1 to 800 mg, from 0.1 to 750 mg, from 0.1 to 700 mg, from 0.1 to 650 mg, from 0.1 to 600 mg, from 0.1 to 550 mg, from 0.1 to 500 mg, from 0.1 to 450 mg, from 0.1 to 400 mg, from 0.1 to 350 mg, from 0.1 to 300 mg, from 0.1 to 250 mg, from 0.1 to 200 mg, from 0.1 to 150 mg, from 0.1 to 100 mg, from 0.1 to 95 mg, from 0.1 to 90 mg, from 0.1 to 85 mg, from 0.1 to 80 mg, from 0.1 to 75 mg, from 0.1 to 70 mg, from 0.1 to 65 mg, from 0.1 to 60 mg, from 0.1 to 55 mg, from 0.1 to 50 mg, from 0.1 to 45 mg, from 0.1 to
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of from about 1 to about 400 mg, from about 2 to about 400 mg, from about 3 to about 400 mg, from about 4 to about 400 mg, from about 5 to about 400 mg, from about 6 to about 400 mg, from about 7 to about 400 mg, from about 8 to about 400 mg, from about 9 to about 400 mg, from about 10 to about 400 mg, from about 20 to about 400 mg, from about 30 to about 400 mg, from about 40 to about 400 mg, from about 50 to about 400 mg, from about 60 to about 400 mg, from about 70 to about 400 mg, from about 80 to about 400 mg, from about 90 to about 400 mg, from about 100 to about 400 mg, from about 110 to about 400 mg, from about 120 to about 400 mg, from about 130 to about 400 mg, from about140 to about 400 mg, from about 150 to about 400 mg, from about160 to about 400 mg
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of from 1 to 400 mg, from 2 to 400 mg, from 3 to 400 mg, from 4 to 400 mg, from 5 to 400 mg, from 6 to 400 mg, from 7 to 400 mg, from 8 to 400 mg, from 9 to 400 mg, from 10 to 400 mg, from 20 to 400 mg, from 30 to 400 mg, from 40 to 400 mg, from 50 to 400 mg, from 60 to 400 mg, from 70 to 400 mg, from 80 to 400 mg, from 90 to 400 mg, from 100 to 400 mg, from 110 to 400 mg, from 120 to 400 mg, from 130 to 400 mg, from about140 to 400 mg, from 150 to 400 mg, from about160 to 400 mg, from 170 to 400 mg, from 180 to 400 mg, from 190 to 400 mg, from 200 to 400 mg, from 210 to 400 mg, from 220 to 400 mg, from 230 to 400 mg, from 240 to 400 mg, from 250 to 400 mg, from
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of from about 10 to about 390 mg, from about 10 to about 380 mg, from about 10 to about 370 mg, from about 10 to about 360 mg, from about 10 to about 350 mg, from about 10 to about 340 mg, from about 10 to about 330 mg, from about 10 to about 320 mg, from about 10 to about 310 mg, from about 10 to about 300 mg, from about 10 to about 290 mg, from about 10 to about 280 mg, from about 10 to about 270 mg, from about 10 to about 260 mg, from about 10 to about 250 mg, from about 10 to about 240 mg, from about 10 to about 230 mg, from about 10 to about 220 mg, from about 10 to about 210 mg, from about 10 to about WSGR Docket No.47991-744.601 200 mg, from about 10 to about 190 mg, from about 10 to about 180 mg, from about 10 to about 170
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of from 10 to 390 mg, from 10 to 380 mg, from 10 to 370 mg, from 10 to 360 mg, from 10 to 350 mg, from 10 to 340 mg, from 10 to 330 mg, from 10 to 320 mg, from 10 to 310 mg, from 10 to 300 mg, from 10 to 290 mg, from 10 to 280 mg, from 10 to 270 mg, from 10 to 260 mg, from 10 to 250 mg, from 10 to 240 mg, from 10 to 230 mg, from 10 to 220 mg, from 10 to 210 mg, from 10 to 200 mg, from 10 to 190 mg, from 10 to 180 mg, from 10 to 170 mg, from 10 to 160 mg, from 10 to 150 mg, from 10 to 140 mg, from 10 to 130 mg, from 10 to 120 mg, from 10 to 110 mg, from 10 to 90 mg, from 10 to 80 mg, from 10 to 70 mg, from 10 to 60 mg
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, about 10 mg, about 11 mg, about 12 mg, about 13 mg, about 14 mg, about 15 mg, about 16 mg, about 17 mg, about 18 mg, about 19 mg, about 20 mg, about 21 mg, about 22 mg, about 23 mg, about 24 mg, about 25 mg, about 26 mg, about 27 mg, about 28 mg, about 29 mg, about 30 mg, about 31 mg, about 32 mg, about 33 mg, about 34 mg, about 35 mg, about 36 mg, about 37 mg, about 38 mg, about 39 mg, about 40 mg, about 41 mg, about 42 mg, about 43 mg, about 44 mg, about 45 mg, about 46 mg, about 47 mg, about 48 mg, about 49 mg, about 50 mg, about 51 mg, about 52 mg, about 53 mg, about
  • the method as described herein comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a first dose of 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 13 mg, 14 mg, 15 mg, 16 mg, 17 WSGR Docket No.47991-744.601 mg, 18 mg, 19 mg, 20 mg, 21 mg, 22 mg, 23 mg, 24 mg, 25 mg, 26 mg, 27 mg, 28 mg, 29 mg, 30 mg, 31 mg, 32 mg, 33 mg, 34 mg, 35 mg, 36 mg, 37 mg, 38 mg, 39 mg, 40 mg, 41 mg, 42 mg, 43 mg, 44 mg, 45 mg, 46 mg, 47 mg, 48 mg, 49 mg, 50 mg, 51 mg, 52 mg, 53 mg, 54 mg, 55 mg, 56 mg, 57 mg, 58 mg, 59 mg, 60 mg, 61 mg, 62 mg, 63 mg, 64 mg,
  • a method of treating or preventing a disease or condition characterized by a reduced expression or function of NaV1.1 protein in a human subject in need thereof comprising administering to the human subject a pharmaceutical composition comprising a first dose of an antisense oligomer (ASO), wherein the ASO comprises a sequence with at least 80% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099, thereby treating or preventing the disease or condition in the human subject; wherein the human subject is at most 18 years old.
  • ASO antisense oligomer
  • the ASO comprises a sequence with at least 80% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B, thereby treating or preventing the disease or condition in the human subject; wherein the human subject is at most 18 years old.
  • a method of treating or preventing a disease or condition characterized by a reduced expression or function of NaV1.1 protein in a human subject in need thereof comprising administering to the human subject a pharmaceutical composition comprising a single dose of an antisense oligomer (ASO), wherein the ASO comprises a sequence with at least 80% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099, thereby treating or preventing the disease or condition in the human subject.
  • ASO antisense oligomer
  • the ASO comprises a sequence with at least 80% sequence identity to any one of any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B, thereby treating or preventing the disease or condition in the human subject.
  • the disease or condition is characterized by a reduced expression or function of Na V 1.1 protein in the subject.
  • the disease or condition is Dravet Syndrome.
  • the subject is characterized by having: (i) seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or Atonic (Drop Attacks), and Clonic; (viii)
  • the subject is characterized by having at least one or more of: (i) seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or Atonic (Drop Attacks), and
  • the subject is characterized by having at least two or more of: (i) seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or Atonic (Drop Attacks), and Clonic; and
  • the subject is characterized by having at least three or more of: (i) seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or Atonic (Drop Attacks), and Clonic; and
  • the subject is characterized by having at least four or more of: (i) seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or Atonic (Drop Attacks), and Clonic; and
  • the subject is characterized by having at least five or more of: (i) seizure onset prior WSGR Docket No.47991-744.601 to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic, Tonic
  • the subject is characterized by having at least six or more of: (i) seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or Atonic (Drop Attacks), and Clonic; and
  • the subject is characterized by having at least seven or more of: (i) seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or Atonic (Drop Attacks), and Clonic; and
  • the subject is characterized by having all eight of: (i) seizure onset prior to 12 months of age with recurrent focal motor or hemiconvulsive or generalized tonic-clonic seizures, which are often prolonged and triggered by hyperthermia; (ii) no past history of causal magnetic resonance WSGR Docket No.47991-744.601 imaging lesion; (iii) no other known etiology of any diseases or conditions except Dravet Syndrome; (iv) normal development at seizure onset; (v) a pathogenic variant, or variant of uncertain significance in an SCN1A gene; (vi) at least 2 prior treatments for epilepsy that either had lack of adequate seizure control; (vii) 4 or more convulsive seizures during the 28 days prior to administering, wherein the convulsive seizures is any one selected from Hemiclonic, Focal with Motor Signs, Focal to Bilateral Tonic Clonic Convulsion, Generalized Tonic Clonic Convulsion, Tonic, Tonic or At
  • the subject is additionally characterized by not having one or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or
  • the subject is characterized by not having one or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide,
  • the subject is additionally characterized by not having two or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having three or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having four or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having five or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having six or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having seven or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having eight or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having nine or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having ten or more of the following: (a) one of the following WSGR Docket No.47991-744.601 mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine
  • the subject is additionally characterized by not having eleven or more of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamotrigine, lacosamide, or rufinamide
  • the subject is additionally characterized by not having all of the following: (a) one of the following mutations in the SCN1A gene: Thr226Met, Leu263Val, Val422Leu, Thr1174Ser, Trp1204Arg, Pro1345Ser, Gln1489Lys, Phe1499Leu, Arg1575Cys, Val1611Phe, Leu1624Pro, Arg1648Cys, Leu1649Gln, WSGR Docket No.47991-744.601 Leu1670Trp, Gly1674Arg, and Asp1866Tyr; (b) a known pathogenic mutation in another gene that causes epilepsy, wherein the pathogenic mutation is homozygous in cases of known recessive disease; (c) currently treated with a sodium channel blocker as maintenance treatment and an anticoagulant, wherein the sodium channel blocker is phenytoin, carbamazepine, oxcarbazepine, lamot
  • the subject has pediatric epilepsy, epileptic encephalopathies, refractory myoclonic epilepsy, or severe myoclonic epilepsy in infancy.
  • the subject has myoclonic epilepsies, generalized epilepsy, epilepsy, brain diseases, central nervous system diseases, nervous system diseases, or epileptic syndromes.
  • the methods of treatment as described herein comprise methods of treating or reducing the likelihood of developing a disease or condition, wherein the disease or condition is pediatric epilepsy, epileptic encephalopathies, refractory myoclonic epilepsy, or severe myoclonic epilepsy in infancy.
  • the methods of treatment as described herein comprise methods of treating or reducing the likelihood of developing a disease or condition, wherein the disease or condition is myoclonic epilepsies, generalized epilepsy, epilepsy, brain diseases, central nervous system diseases, nervous system diseases, or epileptic syndromes.
  • the subject has seizures that are not controlled by current antiepileptic drug (AED) regimen.
  • AED regimen comprises clobazam, cannabidiol, levetiracetam, stiripentol, or valproic acid/valproate.
  • magnetic WSGR Docket No.47991-744.601 resonance imaging refers to a form of medical imaging that measures the response of the atomic nuclei of body tissues to high-frequency radio waves when placed in a strong magnetic field, and that produces images of the internal organs.
  • ketogenic diet refers to a high-fat, adequate-protein, low- carbohydrate diet that in medicine is used, for example, to treat refractory epilepsy in children. The diet forces the body to burn fats rather than carbohydrates.
  • vagal nerve stimulator or “vagus nerve stimulation (VNS)” as used herein, refers to a medical treatment that involves delivering electrical impulses to the vagus nerve. It is, for example, used as an add-on treatment for certain types of intractable epilepsy and treatment-resistant depression.
  • cannabinoid refers to a chemical found in cannabis. Exemplary cannabinoids include, but are not limited to, the phytocannabinoid tetrahydrocannabinol (THC) (Delta9- THC or Delta8-THC), and cannabidiol (CBD). Cannabinoids, as used herein, may be natural or synthetic chemicals.
  • An exemplary main psychoactive component of cannabis is tetrahydrocannabinol (THC).
  • THC tetrahydrocannabinol
  • sodium channel blocker refers to a drug which impair the conduction of sodium ions (Na+) through sodium channels.
  • sodium channel blockers examples include, but are not limited to, alkaloids (e.g., saxitoxin, neosaxitoxin, tetrodotoxin), local anesthetics (e.g., lidocaine), anticonvulsants (e.g., phenytoin, oxcarbazepine (derivative of carbamazepine)), and class Ia (e.g., quinidine, procainamide and disopyramide), class Ib (e.g., lidocaine, mexiletine, tocainide, and phenytoin) and class Ic (e.g., encainide, flecainide, moricizine, and propafenone) antiarrhythmic agents.
  • alkaloids e.g., saxitoxin, neosaxitoxin, tetrodotoxin
  • local anesthetics e.g., lidocaine
  • CSF Cerebrospinal fluid
  • aCSF artificial cerebrospinal fluid
  • CSF drainage shunt refers to a system that drains excess fluid from the brain to another part of the body where the fluid is absorbed as part of the circulatory process. CSF shunts are, for example, used to treat hydrocephalus.
  • EKG ECG
  • ECG electrocardiogram
  • AST Aspartate transaminase
  • AspAT/ASAT/AAT Aspartate aminotransferase
  • GAT AspAT/ASAT/AAT
  • PGP phosphate-dependent transaminase enzyme
  • AST includes any of the recombinant or naturally occurring forms of AST protein or variants or homologs thereof that maintain AST activity, (e.g., within at least 50%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% activity compared to AST).
  • Exemplary AST activity includes, but are not limited to, playing a role in amino acid metabolism, for example, by catalyzing the reversible transfer of an ⁇ -amino group between aspartate and glutamate and, as such.
  • the variants or homologs have at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% amino acid sequence identity across the whole sequence or a portion of the sequence (e.g., a 50, 100, 150 or 200 continuous amino acid portion) compared to a naturally occurring AST protein.
  • the AST protein is substantially identical to the protein identified by the UniProt reference number P17174 or a variant or homolog having substantial identity thereto.
  • the AST protein is substantially identical to the protein identified by the UniProt reference number P00505 or a variant or homolog having substantial identity thereto.
  • ALT also known as alanine aminotransferase (ALAT), serum glutamate-pyruvate transaminase (SGPT), or serum glutamic-pyruvic transaminase (SGPT), as used herein, refers to a transaminase enzyme (EC 2.6.1.2).
  • ALT includes any of the recombinant or naturally occurring forms of ALT protein or variants or homologs thereof that maintain ALT activity, (e.g., within at least 50%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% activity compared to ALT).
  • Exemplary ALT activity includes, but are not limited to, catalyzing the two parts of the alanine cycle.
  • the variants or homologs have at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% amino acid sequence identity across the whole sequence or a portion of the sequence (e.g., a 50, 100, 150 or 200 continuous amino acid portion) compared to a naturally occurring ALT protein.
  • the ALT protein is substantially identical to the protein identified by the UniProt reference number P24298 or a variant or homolog having substantial identity thereto.
  • serum AST level, serum ALT level, and their ratio are measured clinically as biomarkers for liver health.
  • the term “laboratory vale,” as used herein refers to the value obtained by laboratory tests or measurements. Exemplary, non-limiting laboratory tests or measurements may be related to hematology, coagulation, clinical chemistry, plasma, urinalysis, serum, serum or urine pregnancy, urine, or cerebrospinal fluid.
  • the pharmaceutical composition is administered into the intrathecal space of the human subject. In some embodiments, the pharmaceutical composition is administered into the cerebrospinal fluid of the human subject. In some embodiments, the pharmaceutical composition is administered into the brain of the human subject. In some embodiments, the pharmaceutical composition is administered into the cerebrospinal fluid in the brain of the human subject.
  • the pharmaceutical composition is administered as a bolus injection.
  • the method comprises administering the pharmaceutical composition as a bolus injection over 1 to 180 minutes, 175 minutes, 1 to 170 minutes, 1 to 165 minutes, 1 to 160 minutes, 1 to 155 minutes, 1 to 150 minutes, 1 to 145 minutes, 1 to 140 minutes, 1 to 135 minutes, 1 to 130 minutes, 1 WSGR Docket No.47991-744.601 to 125 minutes, 1 to 120 minutes, 1 to 115 minutes, 1 to 110 minutes, 1 to 105 minutes, 1 to 100 minutes, 1 to 95 minutes, 1 to 90 minutes, 1 to 85 minutes, 1 to 80 minutes, 1 to 75 minutes, 1 to 70 minutes, 1 to 65 minutes, 1 to 60 minutes, 1 to 55 minutes, 1 to 50 minutes, 1 to 45 minutes, 1 to 40 minutes, 1 to 35 minutes, 1 to 30 minutes, 1 to 25 minutes, 1 to 20 minutes, 1 to 15 minutes, 1 to 10 minutes, 1 to 5 minutes, or 1 to 3 minutes.
  • the method comprises administering the pharmaceutical composition as a bolus injection using a spinal anesthesia needle.
  • the pharmaceutical composition is administered by infusion with a delivery pump.
  • the pharmaceutical composition is administered by intracerebroventricular injection.
  • the pharmaceutical composition is administered by intrathecal injection.
  • Dravet Syndrome and other related diseases [0506] The terms “condition,” “diseases,” and “disorders” are used herein interchangeably in its broadest sense and include susceptibilities.
  • the disease or condition is Dravet Syndrome.
  • the method reduces or ameliorates at least one symptom of Dravet Syndrome in the human subject.
  • Dravet syndrome otherwise known as severe myoclonic epilepsy of infancy (SMEI)
  • SMEI severe myoclonic epilepsy of infancy
  • Dravet syndrome is an epileptic encephalopathy presenting in the first year of life. Dravet syndrome is an increasingly recognized epileptic encephalopathy in which the clinical diagnosis is supported by the finding of sodium channel gene mutations in approximately 70-80% of patients.
  • DS is a severe and progressive developmental and epileptic encephalopathy that is characterized by high seizure frequency and severity, intellectual disability, and a high risk of sudden unexpected death in epilepsy.
  • VGSCs Voltage-gated sodium channels
  • SCN1A sodium channel ⁇ subunit gene
  • the SCN1A gene is located in the cluster of sodium channel genes on human chromosome 2q24 and encodes the ⁇ -pore forming subunits known as NaV1.1 of the neuronal voltage gated sodium channel.
  • the SCN1A gene spans approximately 100 kb of genomic DNA and comprises 26 exons.
  • the NaV1.1 protein consists of four domains, each with six-transmembrane segments.
  • variants or homologs have at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% nucleotide sequence identity across the whole sequence or a portion of the sequence (e.g., a 50, 100, 150 or 200 continuous nucleotide portion) compared to a naturally occurring SCN1A gene.
  • the SCN1A gene is substantially identical to the gene identified by the Ensembl reference number ENSG00000144285 or a variant or homolog having substantial identity thereto.
  • Alternative splicing events in SCN1A gene can lead to non-productive mRNA transcripts which in turn can lead to aberrant protein expression, and therapeutic agents which can target the alternative splicing events in SCN1A gene can modulate the expression level of functional proteins in DS patients and/or inhibit aberrant protein expression. Such therapeutic agents can be used to treat a condition caused by Na V 1.1 protein deficiency.
  • compositions and methods include antisense oligomers (ASOs) that can cause exon skipping and promote constitutive splicing of SCN1A pre-mRNA.
  • ASOs antisense oligomers
  • functional Na V 1.1 protein can be increased using the methods of the disclosure to treat a condition caused by Na V 1.1 protein deficiency.
  • the disease or condition is induced by a mutation in Na V 1.1 (a protein encoded by the SCN1A gene).
  • Na V 1.1 also known as the sodium channel, voltage-gated, type I, alpha subunit (SCN1A), as used herein, refers to a protein which in humans is encoded by the SCN1A gene.
  • Na V 1.1 includes any of the recombinant or naturally occurring forms of Na V 1.1 protein or variants or homologs thereof that maintain NaV1.1 activity, (e.g., within at least 50%, 80%, 90%, 95%, 96%, 97%, 98%, 99% or 100% activity compared to NaV1.1).
  • the variants or homologs have at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% amino acid sequence identity across the whole sequence or a portion of the sequence (e.g., a 50, 100, 150 or 200 continuous amino acid portion) compared to a naturally occurring NaV1.1 protein.
  • the NaV1.1 protein is substantially identical to the protein identified by the UniProt reference number P35498 or a variant or homolog having substantial identity thereto.
  • the mutation is a loss-of-function mutation in NaV1.1.
  • the loss- of-function mutation in NaV1.1 comprises one or more mutations that decreases or impairs the function WSGR Docket No.47991-744.601 of NaV1.1 (e.g., by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or more) relative to the function of a wild-type NaV1.1.
  • the loss-of-function mutation in NaV1.1 comprises one or more mutations that result in a disease phenotype.
  • Exemplary loss-of-function mutations include, but are not limited to, R859C, T875M, V1353L, I1656M, R1657C, A1685V, M1841T, and R1916G.
  • the mutation is a gain-of-function mutation in NaV1.1.
  • the gain- of-function mutation comprises one or more mutations that prolongs activation of NaV1.1 (e.g., by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or more) relative to the function of a wild-type NaV1.1.
  • the gain-of-function mutation in NaV1.1 comprises one or more mutations that result in a disease phenotype.
  • Exemplary gain-of-function mutations include, but are not limited to, D188V, W1204R, R1648H, and D1866Y.
  • the disease or condition is an encephalopathy.
  • the encephalopathy is induced by a loss-of-function mutation in Na V 1.1.
  • the diseases or conditions are induced by a loss-of-function mutation in Na V 1.1 include, but are not limited to, Dravet Syndrome (DS) (also known as SMEI).
  • DS Dravet Syndrome
  • the present disclosure provides a method of treating or preventing the likelihood of developing a disease or condition in a subject in need thereof, the method comprising administering to the subject the pharmaceutical composition comprising the ASO described herein to decrease the expression of a protein or functional RNA.
  • an ASO is used to decrease the expression of Na V 1.1 protein in cells of a subject having an NMD-inducing exon (NIE) containing pre-mRNA encoding Na V 1.1 protein.
  • NAE NMD-inducing exon
  • the subject has a gain-of-function mutation in Na V 1.1, e.g., migraine.
  • an ASO is used to decrease the expression of Na V 1.1 protein in cells of a subject, the subject has a gain-of-function mutation in Na V 1.1, e.g., migraine, familial hemiplegic, 3.
  • the level of mRNA encoding Na V 1.1 protein is decreased 1.1 to 10-fold, when compared to the amount of mRNA encoding Na V 1.1 protein that is produced in a control cell, e.g., one that is not treated with the ASO or one that is treated with an ASO that does not bind to the targeted portion of the SCN1A NIE containing pre-mRNA.
  • the disease or condition is a Na V 1.1 genetic epilepsy.
  • the Na V 1.1 genetic epilepsy can include a loss-of-function mutation in Na V 1.1 or a gain-of-function mutation in Na V 1.1.
  • the NaV1.1 genetic epilepsy includes one or more hereditary mutations.
  • the NaV1.1 genetic epilepsy includes one or more de novo mutations.
  • the NaV1.1 genetic epilepsy includes Dravet Syndrome (DS) (also known as severe myoclonic epilepsy of infancy or SMEI); severe myoclonic epilepsy of infancy (SMEI)-borderland (SMEB); Febrile seizure (FS); epilepsy, generalized, with febrile seizures plus (GEFS+); epileptic encephalopathy, early infantile, 13; cryptogenic generalized epilepsy; cryptogenic focal epilepsy; myoclonic-astatic epilepsy; Lennox- Gastaut syndrome; West syndrome; idiopathic spasms; early myoclonic encephalopathy; progressive myoclonic epilepsy; alternating hemiplegia of childhood; unclassified epileptic encephalopathy; early infantile SCN1A encephalopathy; early infantile epileptic encephalopathy (EIEE); sudden unexpected WSGR Docket No.47
  • DS Dr
  • the NaV1.1 genetic epilepsy associated with a loss-of-function mutation in NaV1.1 includes Dravet Syndrome (DS) (also known as severe myoclonic epilepsy of infancy or SMEI); severe myoclonic epilepsy of infancy (SMEI)-borderland (SMEB); Febrile seizure (FS); epilepsy, generalized, with febrile seizures plus (GEFS+); epileptic encephalopathy, early infantile, 13; cryptogenic generalized epilepsy; cryptogenic focal epilepsy; myoclonic-astatic epilepsy; Lennox-Gastaut syndrome; West syndrome; idiopathic spasms; early myoclonic encephalopathy; progressive myoclonic epilepsy; alternating hemiplegia of childhood; unclassified epileptic encephalopathy; early infantile SCN1A encephalopathy; early infantile epileptic encephalopathy (EIEE); sudden unexpected death in epilepsy (SUDEP); malignant migrating partial
  • the disease or condition is associated with a haploinsufficiency of the SCN1A gene.
  • diseases or conditions associated with a haploinsufficiency of the SCN1A gene include, but are not limited to, Dravet Syndrome (DS) (also known as SMEI); severe myoclonic epilepsy of infancy (SMEI)-borderland (SMEB); Febrile seizure (FS); epilepsy, generalized, with febrile seizures plus (GEFS+); epileptic encephalopathy, early infantile, 13; cryptogenic generalized epilepsy; cryptogenic focal epilepsy; myoclonic-astatic epilepsy; Lennox-Gastaut syndrome; West syndrome; idiopathic spasms; early myoclonic encephalopathy; progressive myoclonic epilepsy; alternating hemiplegia of childhood; unclassified epileptic encephalopathy; sudden unexpected death in epilepsy (SUDEP); sick sinus syndrome 1; early infantile S
  • DS Dravet
  • the disease or condition is Dravet Syndrome (DS) (also known as SMEI); severe myoclonic epilepsy of infancy (SMEI)-borderland (SMEB); Febrile seizure (FS); epilepsy, generalized, with febrile seizures plus (GEFS+); epileptic encephalopathy, early infantile, 13; cryptogenic generalized epilepsy; cryptogenic focal epilepsy; myoclonic-astatic epilepsy; Lennox-Gastaut syndrome; West syndrome; idiopathic spasms; early myoclonic encephalopathy; progressive myoclonic epilepsy; alternating hemiplegia of childhood; unclassified epileptic encephalopathy; sudden unexpected death in epilepsy (SUDEP); sick sinus syndrome 1; early infantile SCN1A encephalopathy; early infantile epileptic encephalopathy (EIEE); or malignant migrating partial seizures of infancy.
  • DS Dravet Syndrome
  • SMEI severe myoclonic epilepsy
  • the disease or condition is Dravet Syndrome (DS).
  • DS Dravet Syndrome
  • epilepsy refers to a group of neurological disorders characterized by recurrent epileptic seizures.
  • Epileptic seizures refer to episodes that may vary from brief and nearly undetectable periods to long periods of vigorous shaking.
  • Exemplary types of seizure include, but are not limited to, convulsive, non-convulsive, focal, and generalized seizures.
  • Exemplary types of generalized seizures include, but are not limited to, tonic-clonic, tonic, clonic, myoclonic, absence, and atonic seizures.
  • the disease or condition is induced by a gain-of-function mutation in NaV1.1.
  • Exemplary diseases or conditions associated with a gain-of-function mutation in NaV1.1 include, WSGR Docket No.47991-744.601 but are not limited to, migraine.
  • the disease or condition induced by a gain-of-function mutation in NaV1.1 is migraine.
  • the migraine is migraine, familial hemiplegic, 3.
  • the method is a method of decreasing the expression of the NaV1.1 protein by cells of a subject having a NIE containing pre-mRNA encoding the NaV1.1 protein, and wherein the subject has a gain-of-function mutation in NaV1.1.
  • the subject has an allele from which the NaV1.1 protein is produced in an elevated amount or an allele encoding a mutant SCN1A that induces increased activity of NaV1.1 in the cell.
  • the increased activity of NaV1.1 is characterized by a prolonged or near persistent sodium current mediated by the mutant NaV1.1 channel, a slowing of fast inactivation, a positive shift in steady-state inactivation, higher channel availability during repetitive stimulation, increased non-inactivated depolarization-induced persistent sodium currents, delayed entry into inactivation, accelerated recovery from fast inactivation, and/or rescue of folding defects by incubation at lower temperature or co-expression of interacting proteins.
  • the ASO binds to a targeted portion of the NIE containing pre-mRNA transcribed from the second allele, thereby inhibiting or blocking exon skipping of the pseudo-exon from the pre-mRNA, and causing a decrease in the level of mature mRNA encoding functional Na V 1.1 protein, and a decrease in the expression of the Na V 1.1 protein in the cells of the subject.
  • the method reduces or ameliorates at least one symptom of Dravet Syndrome in the human subject.
  • the symptom of Dravet Syndrome is a seizure.
  • the administration reduces or ameliorates seizure frequency, seizure intensity, or seizure duration.
  • the method further comprises assessing tolerability or effectiveness of the pharmaceutical composition.
  • Therapeutic Schedule [0529]
  • the first dose is a single dose. In some embodiments, the first dose is the first of multiple doses.
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from about 0.5 milligrams to about 500 milligrams.
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of 0.1, 0.5, 1, 2.5, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, or 200 mg.
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from about 0.1 to about 1000 mg, from about 0.2 to about 1000 mg, from about 0.3 to about 1000 mg, from about 0.4 to about 1000 mg, from about 0.5 to about 1000 mg, from about 0.6 to about 1000 WSGR Docket No.47991-744.601 mg, from about 0.7 to about 1000 mg, from about 0.8 to about 1000 mg, from about 0.9 to about 1000 mg, 1 to about 1000 mg, from about 2 to about 1000 mg, from about 3 to about 1000 mg, from about 4 to about 1000 mg, from about 5 to about 1000 mg, from about 6 to about 1000 mg, from about 7 to about 1000 mg, from about 8 to about 1000 mg, from about 9 to about 1000 mg, from about 10 to about 1000 mg, from about 15 to about 1000 mg, from about 20 to about 1000 mg, from about 25 to about 1000 mg, from about 30 to about 1000 mg, from about 35 to about 1000 mg
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from 0.1 to 1000 mg, from 0.2 to 1000 mg, from 0.3 to 1000 mg, from 0.4 to 1000 mg, from 0.5 to 1000 mg, from 0.6 to 1000 mg, from 0.7 to 1000 mg, from 0.8 to 1000 mg, from 0.9 to 1000 mg, 1 to 1000 mg, from 2 to 1000 mg, from 3 to 1000 mg, from 4 to 1000 mg, from 5 to 1000 mg, from 6 to 1000 mg, from 7 to 1000 mg, from 8 to 1000 mg, from 9 to 1000 mg, from 10 to 1000 mg, from 15 to 1000 mg, from 20 to 1000 mg, from 25 to 1000 mg, from 30 to 1000 mg, from 35 to 1000 mg, from 40 to 1000 mg, from 45 to 1000 mg, from 50 to 1000 mg, from 55 to 1000 mg, from 60 to 1000 mg, from 65 to 1000 mg, from 70 to 1000 mg, from 75 to 1000 mg, from 80 to 1000 mg, from 85 to 1000 mg, from
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from about 0.1 to about 950 mg, from about 0.1 to about 900 mg, from about 0.1 to about 850 mg, from about 0.1 to about 800 mg, from about 0.1 to about 750 mg, from about 0.1 to about 700 mg, from about 0.1 to about 650 mg, from about 0.1 to about 600 mg, from about 0.1 to about 550 mg, from about 0.1 to about 500 mg, from about 0.1 to about 450 mg, from about 0.1 to about 400 mg, from about 0.1 to about 350 mg, from about 0.1 to about 300 mg, from about 0.1 to about 250 mg, from about 0.1 to about 200 mg, from about 0.1 to about 150 mg, from about 0.1 to about 100 mg, from about 0.1 to about WSGR Docket No.47991-744.601 95 mg, from about 0.1 to about 90 mg, from about 0.1 to about 85 mg, from about
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from 0.1 to 950 mg, from 0.1 to 900 mg, from 0.1 to 850 mg, from 0.1 to 800 mg, from 0.1 to 750 mg, from 0.1 to 700 mg, from 0.1 to 650 mg, from 0.1 to 600 mg, from 0.1 to 550 mg, from 0.1 to 500 mg, from 0.1 to 450 mg, from 0.1 to 400 mg, from 0.1 to 350 mg, from 0.1 to 300 mg, from 0.1 to 250 mg, from 0.1 to 200 mg, from 0.1 to 150 mg, from 0.1 to 100 mg, from 0.1 to 95 mg, from 0.1 to 90 mg, from 0.1 to 85 mg, from 0.1 to 80 mg, from 0.1 to 75 mg, from 0.1 to 70 mg, from 0.1 to 65 mg, from 0.1 to 60 mg, from 0.1 to 55 mg, from 0.1 to 50 mg, from 0.1 to 45 mg, from 0.1
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from about 1 to about 400 mg, from about 2 to about 400 mg, from about 3 to about 400 mg, from about 4 to about 400 mg, from about 5 to about 400 mg, from about 6 to about 400 mg, from about 7 to about 400 mg, from about 8 to about 400 mg, from about 9 to about 400 mg, from about 10 to about 400 mg, from about 20 to about 400 mg, from about 30 to about 400 mg, from about 40 to about 400 mg, from about 50 to about 400 mg, from about 60 to about 400 mg, from about 70 to about 400 mg, from about 80 to about 400 mg, from about 90 to about 400 mg, from about 100 to about 400 mg, from about 110 to about 400 mg, from about 120 to about 400 mg, from about 130 to about 400 mg, from about140 to about 400 mg, from about 150 to about 400 mg, from about160 to about 400 mg, from about 170 to about 400 mg, from about 180 to
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from 1 to 400 mg, from 2 to 400 mg, from 3 to 400 mg, from 4 to 400 mg, from 5 to 400 mg, from 6 to 400 mg, from 7 to 400 mg, from 8 to 400 mg, from 9 to 400 mg, from 10 to 400 mg, from 20 to 400 mg, from 30 to 400 mg, from 40 to 400 mg, from 50 to 400 mg, from 60 to 400 mg, from 70 to 400 mg, from 80 to 400 mg, from 90 to 400 mg, from 100 to 400 mg, from 110 to 400 mg, from 120 to 400 mg, from 130 to 400 mg, from about140 to 400 mg, from 150 to 400 mg, from about160 to 400 mg, from 170 to 400 mg, from 180 to 400 mg, from 190 to 400 mg, from 200 to 400 mg, from 210 to 400 mg, from 220 to 400 mg, from 230 to 400 mg, from 240 to 400 mg, from 250 to 400 mg, from
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from about 10 to about 390 mg, from about 10 to about 380 mg, from about 10 to about 370 mg, from about 10 to about 360 mg, from about 10 to about 350 mg, from about 10 to about 340 mg, from about 10 to about 330 mg, from about 10 to about 320 mg, from about 10 to about 310 mg, from about 10 to about 300 mg, from about 10 to about 290 mg, from about 10 to about 280 mg, from about 10 to about 270 mg, from about 10 to about 260 mg, from about 10 to about 250 mg, from about 10 to about 240 mg, from about 10 to about 230 mg, from about 10 to about 220 mg, from about 10 to about 210 mg, from about 10 to about 200 mg, from about 10 to about 190 mg, from about 10 to about 180 mg, from about 10 to about 170 mg, from about 10 to about 160 mg, from about 10 to
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of from 10 to 390 mg, from 10 to 380 mg, from 10 to 370 mg, from 10 to 360 mg, from 10 to 350 mg, from 10 to 340 mg, from 10 to 330 mg, from 10 to 320 mg, from 10 to 310 mg, from 10 to 300 mg, from 10 to 290 mg, from 10 to 280 mg, from 10 to 270 mg, from 10 to 260 mg, from 10 to 250 mg, from 10 to 240 mg, from 10 to 230 mg, from 10 to 220 mg, from 10 to 210 mg, from 10 to 200 mg, from 10 to 190 mg, from 10 to 180 mg, from 10 to 170 mg, from 10 to 160 mg, from 10 to 150 mg, from 10 to 140 mg, from 10 to 130 mg, from 10 to 120 mg, from 10 to 110 mg, from 10 to 90 mg, from 10 to 80 mg, WSGR Docket No.4799
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, about 10 mg, about 11 mg, about 12 mg, about 13 mg, about 14 mg, about 15 mg, about 16 mg, about 17 mg, about 18 mg, about 19 mg, about 20 mg, about 21 mg, about 22 mg, about 23 mg, about 24 mg, about 25 mg, about 26 mg, about 27 mg, about 28 mg, about 29 mg, about 30 mg, about 31 mg, about 32 mg, about 33 mg, about 34 mg, about 35 mg, about 36 mg, about 37 mg, about 38 mg, about 39 mg, about 40 mg, about 41 mg, about 42 mg, about 43 mg, about 44 mg, about 45 mg, about 46 mg, about 47 mg, about 48 mg, about 49 mg, about 50 mg, about 51 mg, about 52 mg, about 53 mg,
  • the method as described herein further comprises administering to the human subject a pharmaceutical composition comprising the ASO as described herein at a subsequent dose of 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 13 mg, 14 mg, 15 mg, 16 mg, 17 mg, 18 mg, 19 mg, 20 mg, 21 mg, 22 mg, 23 mg, 24 mg, 25 mg, 26 mg, 27 mg, 28 mg, 29 mg, 30 mg, 31 mg, 32 mg, 33 mg, 34 mg, 35 mg, 36 mg, 37 mg, 38 mg, 39 mg, 40 mg, 41 mg, 42 mg, 43 mg, 44 mg, 45 mg, 46 mg, 47 mg, 48 mg, 49 mg, 50 mg, 51 mg, 52 mg, 53 mg, 54 mg, 55 mg, 56 mg, 57 mg, 58 mg, 59 mg, 60 mg, 61 mg, 62 mg, 63 mg, 64 mg, 65 mg, 66 mg, 67 mg, 68 mg
  • the method as described herein further comprises assessing tolerability or effectiveness of the pharmaceutical composition.
  • the subsequent dose is lower than the previous dose following an indication that administration of the previous dose is not tolerated.
  • the subsequent dose is the same as the previous dose following an indication that administration of the previous dose is tolerated.
  • the subsequent dose is higher than the previous dose following an indication that administration of the previous dose is tolerated.
  • the subsequent dose is the same as the previous dose following an indication that administration of the previous dose is effective.
  • the subsequent dose is lower than the previous dose following an indication that administration of the previous dose is effective.
  • the subsequent dose is higher than the previous dose following an indication that administration of the previous dose is not effective. [0544] In some embodiments, the subsequent dose is lower than the previous dose following an indication that administration of the previous dose is not tolerated. [0545] In some embodiments, the subsequent dose is the same as the previous dose following an indication that administration of the previous dose is tolerated. [0546] In some embodiments, the subsequent dose is higher than the previous dose following an indication that administration of the previous dose is tolerated. WSGR Docket No.47991-744.601 [0547] In some embodiments, the subsequent dose is the same as the previous dose following an indication that administration of the previous dose is effective.
  • the subsequent dose is lower than the previous dose following an indication that administration of the previous dose is effective.
  • the subsequent dose is higher than the previous dose following an indication that administration of the previous dose is not effective.
  • the subsequent doses are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months after administration of the previous dose.
  • the subsequent doses are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 hours after administration of the previous dose.
  • the subsequent doses are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 days after administration of the previous dose. In some embodiments, the subsequent doses are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 36 weeks after administration of the previous dose. In some embodiments, the subsequent doses are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 36 months after administration of the previous dose.
  • the subsequent doses are administered at least 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 years after administration of the previous dose.
  • the subsequent doses are administered at the same interval. For example, every subsequent dose is administered at the interval of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 hours after administration of the previous dose. In some embodiments, every subsequent dose is administered at the interval of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 days after administration of the previous dose.
  • every subsequent dose is administered at the interval of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 36 weeks after administration of the previous dose. In some embodiments, every subsequent dose is administered at the interval of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 36 months after administration of the previous dose. [0553] In some embodiments, the subsequent doses are administered at the different intervals. [0554] In some embodiments, the dose frequency is maintained or reduced following an indication that the previous dose is effective.
  • the dose frequency is increased following an indication that the previous dose is not effective.
  • the method further comprises administrating at least one additional therapeutic agent or therapy.
  • the at least one additional therapeutic agent or therapy is administered at the same time as the dose.
  • the at least one additional therapeutic agent or therapy is administered prior to WSGR Docket No.47991-744.601 administration of the dose.
  • the at least one additional therapeutic agent or therapy is administered after administration of the dose.
  • the reduced expression or function of NaV1.1 protein is associated with an altered splicing of a nonsense-mediated RNA decay-inducing exon from a pre-mRNA that contains the NMD exon and encodes NaV1.1 protein.
  • the ASO promotes exclusion of the NMD exon from the pre-mRNA that contains the NMD exon and that encodes the NaV1.1 protein.
  • the ASO binds to a targeted portion of a pre-mRNA that contains the NMD exon and that encodes the NaV1.1 protein.
  • Compositions [0558] In some embodiments, the ASO comprises a sequence with at least 83%, 88%, 94% or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the ASO consists of a sequence with at least 83%, 88%, 94% or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099. In some embodiments, the ASO comprises a sequence with at least 83%, 88%, 94% or 100% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B. In some embodiments, the ASO consists of a sequence with at least 83%, 88%, 94% or 100% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the ASO comprises a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the ASO consists of a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the ASO comprises a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the ASO consists of a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the ASO as described herein comprises at least one modified sugar moiety.
  • the ASO as described herein comprises T-methoxyethyl sugar moiety.
  • the T-methoxyethyl sugar moiety is a T-2’-methoxyethyl sugar moiety.
  • the ASO as described herein comprises a 2’-O-methoxyethyl moiety.
  • the ASO as described herein comprises a thymidine comprising a 2’-O-methoxyethyl moiety.
  • each nucleobase of the ASO as described herein comprises a 2’-O- methoxyethyl moiety.
  • the ASO as described herein consists of from 8 to 50 nucleobases. In some embodiments, the ASO as described herein consists of from 16 to 20 nucleobases. In some embodiments, the ASO as described herein consists of from 12 to 20 nucleobases. In some embodiments, the ASO as described herein consists of from 8 to 20 nucleobases.
  • the ASO as described herein consists of from 5 to 100, from 6 to 100, from 7 to 100, from 8 to 100, from 9 to 100, from 10 to 100, from 11 to 100, from 12 to 100, from 13 to 100, from 14 to 100, from 15 to 100, from 16 to 100, from 17 to 100, from 18 to 100, from 19 to 100, from 20 to 100, from 21 to 100, from 22 to 100, from 23 to 100, from 24 to 100, from 25 to 100, from 30 to 100, from 35 to 100, from 40 to 100, from 45 to 100, from 50 to 100, from 55 to 100, from 60 to 100, from 65 to 100, from 70 to 100, from 75 to 100, from 80 to 100, from 85 to 100, or from 90 to 100 nucleobases.
  • the ASO as described herein consists of from 5 to 100, 5 to 95, 5 to 90, 5 to 85, 5 to 80, 5 to 75, 5 to 70, 5 to 65, 5 to 60, 5 to 55, 5 to 50, 5 to 45, 5 to 40, 5 to 35, 5 to 30, 5 to 25, 5 to 20, 5 to 15, or 5 to 10 nucleobases.
  • the ASO as described herein consists of from 8 to 50, from 8 to 45, from 8 to 40, from 8 to 35, from 8 to 30, from 8 to 29, from 8 to 28, from 8 to 27, from 8 to 26, from 8 to 25, from 8 to 24, from 8 to 23, from 8 to 22, from 8 to 21, from 8 to 20, from 8 to 19, from 8 to 18, from 8 to 17, or from 8 to 16 nucleobases.
  • the ASO as described herein consists of from 9 to 20, from 10 to 20, from 11 to 20, from 12 to 20, from 13 to 20, from 14 to 20, from 15 to 20, from 16 to 20, from 17 to 20, or from 18 to 20 nucleobases.
  • the ASO as described herein consists of 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, or 40 nucleobases.
  • the ASO as described herein comprises a 5’-methylcytosine (5’-MeC).
  • each cytosine of the ASO as described herein is a 5’-methylcytosine (5’-MeC).
  • the ASO as described herein comprises a phosphorothioate linkage.
  • each internucleoside linkage of the ASO as described herein is a phosphorothioate linkage.
  • the ASO as described herein comprises a locked nucleic acid (LNA).
  • LNA locked nucleic acid
  • the ASO as described herein comprises least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 LNAs.
  • the ASO as described herein comprises 1 to 20, 1 to 19, 1 to 18, 1 to 17, 1 to 16, 1 to 15, 1 to 14, 1 to 13, 1 to 12, 1 to 11, 1 to 10, 1 to 9, 1 to 8, 1 to 7, 1 to 6, 1 to 5, 1 to 4, 1 to 3, or 1 to 2 LNAs.
  • the ASO as described herein comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 LNAs.
  • the 5’ end nucleotide of the ASO as described herein is a LNA.
  • the 3’ end nucleotide of the ASO as described herein is a LNA.
  • the 5’ and 3’ end nucleotides of the ASO as described herein are LNAs.
  • ASO Antisense Oligomers
  • ASOs are designed so that they bind (hybridize) to a target nucleic acid (e.g., a targeted portion of a pre-mRNA transcript) and remain hybridized under physiological conditions. Typically, if they hybridize to a site other than the intended (targeted) nucleic acid sequence, they hybridize to a limited number of sequences that are not a target nucleic acid (to a few sites other than a target nucleic acid).
  • a target nucleic acid e.g., a targeted portion of a pre-mRNA transcript
  • Design of an ASO can take into consideration the occurrence of the nucleic acid sequence of the targeted portion of the pre-mRNA transcript or a sufficiently similar nucleic acid sequence in other locations in the genome or cellular pre-mRNA or transcriptome, such that the likelihood the ASO will bind other sites and cause “off-target” effects is limited.
  • Any antisense oligomers known in the art for example in PCT Application No. PCT/US2014/054151, published as WO 2015/035091, titled “Reducing Nonsense-Mediated mRNA Decay,” incorporated by reference herein, can be used to practice the methods described herein.
  • ASOs “specifically hybridize” to or are “specific” to a target nucleic acid or a targeted portion of a NIE containing pre-mRNA.
  • hybridization occurs with a T m substantially greater than 37 °C, preferably at least 50 °C, and typically between 60 °C, to approximately 90 °C.
  • T m is the temperature at which 50% of a target sequence hybridizes to a complementary oligonucleotide.
  • Oligomers such as oligonucleotides, are “complementary” to one another when hybridization occurs in an antiparallel configuration between two single-stranded polynucleotides.
  • a double-stranded polynucleotide can be “complementary” to another polynucleotide, if hybridization can occur between one of the strands of the first polynucleotide and the second.
  • Complementarity (the degree to which one polynucleotide is complementary with another) is quantifiable in terms of the proportion (e.g., the percentage) of bases in opposing strands that are expected to form hydrogen bonds with each other, according to generally accepted base-pairing rules.
  • ASO antisense oligomer
  • ASOs can comprise at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence complementarity to a target region within the target nucleic acid sequence to which they are targeted.
  • an ASO in which 18 of 20 nucleobases of the oligomeric compound are complementary to a target region, and would therefore specifically hybridize would represent 90 percent complementarity.
  • the remaining non- complementary nucleobases may be clustered together or interspersed with complementary nucleobases and need not be contiguous to each other or to complementary nucleobases.
  • Percent complementarity of WSGR Docket No.47991-744.601 an ASO with a region of a target nucleic acid can be determined routinely using BLAST programs (basic local alignment search tools) and PowerBLAST programs known in the art (Altschul, et al., J. Mol. Biol., 1990, 215, 403-410; Zhang and Madden, Genome Res., 1997, 7, 649-656, of which entire content is incorporated herein by reference).
  • An ASO need not hybridize to all nucleobases in a target sequence and the nucleobases to which it does hybridize may be contiguous or noncontiguous. ASOs may hybridize over one or more segments of a pre-mRNA transcript, such that intervening or adjacent segments are not involved in the hybridization event (e.g., a loop structure or hairpin structure may be formed). In certain embodiments, an ASO hybridizes to noncontiguous nucleobases in a target pre-mRNA transcript. For example, an ASO can hybridize to nucleobases in a pre-mRNA transcript that are separated by one or more nucleobase(s) to which the ASO does not hybridize.
  • the ASOs described herein comprise nucleobases that are complementary to nucleobases present in a targeted portion of a NIE containing pre-mRNA.
  • the term ASO embodies oligonucleotides and any other oligomeric molecule that comprises nucleobases capable of hybridizing to a complementary nucleobase on a target mRNA but does not comprise a sugar moiety, such as a peptide nucleic acid (PNA).
  • PNA peptide nucleic acid
  • the ASOs may comprise naturally occurring nucleotides, nucleotide analogs, modified nucleotides, or any combination of two or three of the preceding.
  • the term “naturally occurring nucleotides” includes deoxyribonucleotides and ribonucleotides.
  • modified nucleotides includes nucleotides with modified or substituted sugar groups and/or having a modified backbone. In some embodiments, all of the nucleotides of the ASO are modified nucleotides.
  • Chemical modifications of ASOs or components of ASOs that are compatible with the methods and compositions described herein will be evident to one of skill in the art and can be found, for example, in U.S. Pat.8,258,109 B2, U.S. Pat. No.5,656,612, U.S. Patent Publication No.2012/0190728, and Dias and Stein, Mol. Cancer Ther.2002, 347-355, herein incorporated by reference in their entirety.
  • nucleobases of an ASO may be any naturally occurring, unmodified nucleobase such as adenine, guanine, cytosine, thymine and uracil, or any synthetic or modified nucleobase that is sufficiently similar to an unmodified nucleobase such that it is capable of hydrogen bonding with a nucleobase present on a target pre-mRNA.
  • modified nucleobases include, without limitation, hypoxanthine, xanthine, 7-methylguanine, 5, 6-dihydrouracil, 5-methylcytosine, and 5- hydroxymethoylcytosine.
  • the ASOs described herein also comprise a backbone structure that connects the components of an oligomer.
  • backbone structure and “oligomer linkages” may be used interchangeably and refer to the connection between monomers of the ASO.
  • the backbone comprises a 3'-5' phosphodiester linkage connecting sugar moieties of the oligomer.
  • the backbone structure or oligomer linkages of the ASOs described herein may include (but are not limited to) phosphorothioate, phosphorodithioate, phosphoroselenoate, phosphorodiselenoate, phosphoroanilothioate, phosphoraniladate, phosphoramidate, and the like.
  • the backbone structure of the ASO does not contain phosphorous but rather contains peptide bonds, for example in a peptide nucleic acid (PNA), or linking groups including carbamate, amides, and linear and cyclic hydrocarbon groups.
  • PNA peptide nucleic acid
  • the backbone modification is a phosphothioate linkage.
  • the backbone modification is a phosphoramidate linkage.
  • the stereochemistry at each of the phosphorus internucleotide linkages of the ASO backbone is random.
  • the stereochemistry at each of the phosphorus internucleotide linkages of the ASO backbone is controlled and is not random.
  • U.S. Pat. App. Pub. No. 2014/0194610 “Methods for the Synthesis of Functionalized Nucleic Acids,” incorporated herein by reference, describes methods for independently selecting the handedness of chirality at each phosphorous atom in a nucleic acid oligomer.
  • an ASO used in the methods of the invention including, but not limited to, any of the ASOs set forth herein in Tables 5 and 6, comprises an ASO having phosphorus internucleotide linkages that are not random.
  • a composition used in the methods of the invention comprises a pure diastereomeric ASO.
  • a composition used in the methods of the invention comprises an ASO that has diastereomeric purity of at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, about 100%, about 90% to about 100%, about 91% to about 100%, about 92% to about 100%, about 93% to about 100%, about 94% to about 100%, about 95% to about 100%, about 96% to about 100%, about 97% to about 100%, about 98% to about 100%, or about 99% to about 100%.
  • the ASO has a nonrandom mixture of Rp and Sp configurations at its phosphorus internucleotide linkages.
  • Rp and Sp are required in antisense oligonucleotides or antisense oligomers to achieve a balance between good activity and nuclease stability.
  • an ASO used in the methods of the invention comprises about 5-100% Rp, at least about 5% Rp, at least about 10% Rp, at least about 15% Rp, at least about 20% Rp, at least about 25% Rp, at least about 30% Rp, at least about 35% Rp, at least about 40% Rp, at least about 45% Rp, at least about 50% Rp, at least about 55% Rp, at least about 60% Rp, at least about 65% Rp, at least about 70% Rp, at least about 75% Rp, at least about 80% Rp, at least about 85% Rp, at least about 90% Rp, or at least about 95% Rp, with the remainder Sp, or about 100% Rp.
  • an ASO used in the methods of the invention comprises about 10% to about 100% Rp, about 15% to about 100% Rp, about 20% to about 100% Rp, about 25% to about 100% Rp, about 30% to about WSGR Docket No.47991-744.601 100% Rp, about 35% to about 100% Rp, about 40% to about 100% Rp, about 45% to about 100% Rp, about 50% to about 100% Rp, about 55% to about 100% Rp, about 60% to about 100% Rp, about 65% to about 100% Rp, about 70% to about 100% Rp, about 75% to about 100% Rp, about 80% to about 100% Rp, about 85% to about 100% Rp, about 90% to about 100% Rp, or about 95% to about 100% Rp, about 20% to about 80% Rp, about 25% to about 75% Rp, about 30% to about 70% Rp, about 40% to about 60% Rp, or about 45% to about 55% Rp, about 10% to about 100% Rp, about 15% to about 100% Rp, about 20% to about 100% Rp, about 25% to about 100% R
  • an ASO used in the methods of the invention comprises about 5-100% Sp, at least about 5% Sp, at least about 10% Sp, at least about 15% Sp, at least about 20% Sp, at least about 25% Sp, at least about 30% Sp, at least about 35% Sp, at least about 40% Sp, at least about 45% Sp, at least about 50% Sp, at least about 55% Sp, at least about 60% Sp, at least about 65% Sp, at least about 70% Sp, at least about 75% Sp, at least about 80% Sp, at least about 85% Sp, at least about 90% Sp, or at least about 95% Sp, with the remainder Rp, or about 100% Sp.
  • an ASO used in the methods of the invention comprises about 10% to about 100% Sp, about 15% to about 100% Sp, about 20% to about 100% Sp, about 25% to about 100% Sp, about 30% to about 100% Sp, about 35% to about 100% Sp, about 40% to about 100% Sp, about 45% to about 100% Sp, about 50% to about 100% Sp, about 55% to about 100% Sp, about 60% to about 100% Sp, about 65% to about 100% Sp, about 70% to about 100% Sp, about 75% to about 100% Sp, about 80% to about 100% Sp, about 85% to about 100% Sp, about 90% to about 100% Sp, or about 95% to about 100% Sp, about 20% to about 80% Sp, about 25% to about 75% Sp, about 30% to about 70% Sp, about 40% to about 60% Sp, or about 45% to about 55% Sp, with the remainder Rp.
  • an ASO used in the methods of the invention comprises about 5- 100% Rp, at least about 5% Rp, at least about 10% Rp, at least about 15% Rp, at least about 20% Rp, at least about 25% Rp, at least about 30% Rp, at least about 35% Rp, at least about 40% Rp, at least about 45% Rp, at least about 50% Rp, at least about 55% Rp, at least about 60% Rp, at least about 65% Rp, at least about 70% Rp, at least about 75% Rp, at least about 80% Rp, at least about 85% Rp, at least about 90% Rp, or at least about 95% Rp, with the remainder Sp, or about 100% Rp.
  • an ASO used in the methods of the invention comprises about 10% to about 100% Rp, about 15% to about 100% Rp, about 20% to about 100% Rp, about 25% to about 100% Rp, about 30% to about 100% Rp, about 35% to about 100% Rp, about 40% to about 100% Rp, about 45% to about 100% Rp, about 50% to about 100% Rp, about 55% to about 100% Rp, about 60% to about 100% Rp, about 65% to about 100% Rp, about 70% to about 100% Rp, about 75% to about 100% Rp, about 80% to about 100% Rp, about 85% to about 100% Rp, about 90% to about 100% Rp, or about 95% to about 100% Rp, about 20% to about 80% Rp, about 25% to about 75% Rp, about 30% to about 70% Rp, about 40% to about 60% Rp, or about 45% to about 55% Rp, with the remainder
  • an ASO used in the methods of the invention comprises about 5- 100% Sp, at least about 5% Sp, at least about 10% Sp, at least about 15% Sp, at least about 20% Sp, at least about 25% Sp, at least about 30% Sp, at least about 35% Sp, at least about 40% Sp, at least about 45% Sp, at least about 50% Sp, at least about 55% Sp, at least about 60% Sp, at least about 65% Sp, at least about 70% Sp, at least about 75% Sp, at least about 80% Sp, at least about 85% Sp, at least about 90% Sp, or at least about 95% Sp, with the remainder Rp, or about 100% Sp.
  • an ASO used in the methods of the invention comprises about 10% to about 100% Sp, about 15% to about 100% Sp, about 20% to about 100% Sp, about 25% to about 100% Sp, about 30% to about 100% Sp, about 35% to about 100% Sp, about 40% to about 100% Sp, about 45% to about 100% Sp, about 50% to about 100% Sp, about 55% to about 100% Sp, about 60% to about 100% Sp, about 65% to about 100% Sp, about 70% to about 100% Sp, about 75% to about 100% Sp, about 80% to about 100% Sp, about 85% to about 100% Sp, about 90% to about 100% Sp, or about 95% to about 100% Sp, about 20% to about 80% Sp, about 25% to about 75% Sp, about 30% to about 70% Sp, about 40% to about 60% Sp, or about 45% to about 55% Sp, with the remainder Rp.
  • an ASO used in the methods of the invention comprises about 5-100% Rp, at least about 5% Rp, at least about 10% Rp, at least about 15% Rp, at least about 20% Rp, at least about 25% Rp, at least about 30% Rp, at least about 35% Rp, at least about 40% Rp, at least about 45% Rp, at least about 50% Rp, at least about 55% Rp, at least about 60% Rp, at least about 65% Rp, at least about 70% Rp, at least about 75% Rp, at least about 80% Rp, at least about 85% Rp, at least about 90% Rp, or at least about 95% Rp, with the remainder Sp, or about 100% Rp.
  • an ASO used in the methods of the invention comprises about 10% to about 100% Rp, about 15% to about 100% Rp, about 20% to about 100% Rp, about 25% to about 100% Rp, about 30% to about 100% Rp, about 35% to about 100% Rp, about 40% to about 100% Rp, about 45% to about 100% Rp, about 50% to about 100% Rp, about 55% to about 100% Rp, about 60% to about 100% Rp, about 65% to about 100% Rp, about 70% to about 100% Rp, about 75% to about 100% Rp, about 80% to about 100% Rp, about 85% to about 100% Rp, about 90% to about 100% Rp, or about 95% to about 100% Rp, about 20% to about 80% Rp, about 25% to about 75% Rp, about 30% to about 70% Rp, about 40% to about 10% Rp, about 15% to about 100% Rp, about 20% to about 100% Rp, about 25% to about 100% Rp, about 30% to about 100% Rp, about 35% to about 100% Rp, about 40% to about 100% Rp, about 45% to about 100% R
  • an ASO used in the methods of the invention comprises about 5-100% Sp, at least about 5% Sp, at least about 10% Sp, at least about 15% Sp, at least about 20% Sp, at least about 25% Sp, at least about 30% Sp, at least about 35% Sp, at least about 40% Sp, at least about 45% Sp, at least about 50% Sp, at least about 55% Sp, at least about WSGR Docket No.47991-744.601 60% Sp, at least about 65% Sp, at least about 70% Sp, at least about 75% Sp, at least about 80% Sp, at least about 85% Sp, at least about 90% Sp, or at least about 95% Sp, with the remainder Rp, or about 100% Sp.
  • an ASO used in the methods of the invention comprises about 10% to about 100% Sp, about 15% to about 100% Sp, about 20% to about 100% Sp, about 25% to about 100% Sp, about 30% to about 100% Sp, about 35% to about 100% Sp, about 40% to about 100% Sp, about 45% to about 100% Sp, about 50% to about 100% Sp, about 55% to about 100% Sp, about 60% to about 100% Sp, about 65% to about 100% Sp, about 70% to about 100% Sp, about 75% to about 100% Sp, about 80% to about 100% Sp, about 85% to about 100% Sp, about 90% to about 100% Sp, or about 95% to about 100% Sp, about 20% to about 80% Sp, about 25% to about 75% Sp, about 30% to about 70% Sp, about 40% to about 60% Sp, or about 45% to about 55% Sp, with the remainder Rp.
  • Any of the ASOs described herein may contain a sugar moiety that comprises ribose or deoxyribose, as present in naturally occurring nucleotides, or a modified sugar moiety or sugar analog, including a morpholine ring.
  • Non-limiting examples of modified sugar moieties include 2' substitutions such as 2'-O-methyl (2'-O-Me), 2'-O-methoxyethyl (2'MOE), 2'-O-aminoethyl, 2'F; N3'->P5' phosphoramidate, 2' dimethylaminooxyethoxy, 2' dimethylaminoethoxyethoxy, 2'-guanidinidium, 2'-O- guanidinium ethyl, carbamate modified sugars, and bicyclic modified sugars.
  • the sugar moiety modification is selected from 2'-O-Me, 2'F, and 2'MOE.
  • the sugar moiety modification is an extra bridge bond, such as in a locked nucleic acid (LNA).
  • the sugar analog contains a morpholine ring, such as phosphorodiamidate morpholino (PMO).
  • the sugar moiety comprises a ribofuransyl or 2' deoxyribofuransyl modification.
  • the sugar moiety comprises 2'4'-constrained 2'O-methyloxyethyl (cMOE) modifications.
  • the sugar moiety comprises cEt 2', 4' constrained 2'-O ethyl BNA modifications.
  • the sugar moiety comprises tricycloDNA (tcDNA) modifications.
  • the sugar moiety comprises ethylene nucleic acid (ENA) modifications.
  • the sugar moiety comprises MCE modifications. Modifications are known in the art and described in the literature, e.g., by Jarver, et al., 2014, Nucleic Acid Therapeutics 24(1): 37-47, incorporated by reference for this purpose herein. “A Chemical View of Oligonucleotides for Exon Skipping and Related Drug Applications,” Nucleic Acid Therapeutics 24(1): 37-47, incorporated by reference for this purpose herein.
  • each monomer of the ASO is modified in the same way, for example each linkage of the backbone of the ASO comprises a phosphorothioate linkage or each ribose sugar moiety comprises a 2'O-methyl modification.
  • Such modifications that are present on each of the monomer components of an ASO are referred to as “uniform modifications.”
  • a combination of different modifications may be desired, for example, an ASO may comprise a combination of phosphorodiamidate linkages and sugar moieties comprising morpholine rings (morpholinos).
  • the ASO comprises one or more backbone modifications.
  • the ASO comprises one or more sugar moiety modification.
  • the ASO comprises one or more backbone modifications and one or more sugar moiety modifications.
  • the ASO comprises a 2'MOE modification and a phosphorothioate backbone.
  • the ASO comprises a phosphorodiamidate morpholino (PMO).
  • the ASO comprises a peptide nucleic acid (PNA).
  • any of the ASOs or any component of an ASO may be modified in order to achieve desired properties or activities of the ASO or reduce undesired properties or activities of the ASO.
  • an ASO or one or more components of any ASO may be modified to enhance binding affinity to a target sequence on a pre-mRNA transcript; reduce binding to any non-target sequence; reduce degradation by cellular nucleases (i.e., RNase H); improve uptake of the ASO into a cell and/or into the nucleus of a cell; alter the pharmacokinetics or pharmacodynamics of the ASO; and/or modulate the half-life of the ASO.
  • the ASOs are comprised of 2'-O-(2-methoxyethyl) (MOE) phosphorothioate-modified nucleotides.
  • MOE 2'-O-(2-methoxyethyl)
  • ASOs comprised of such nucleotides are especially well-suited to the methods disclosed herein; oligomers having such modifications have been shown to have significantly enhanced resistance to nuclease degradation and increased bioavailability, making them suitable, for example, for oral delivery in some embodiments described herein.
  • the left-hand end of single-stranded nucleic acid e.g., pre-mRNA transcript, oligonucleotide, ASO, etc.
  • sequences is the 5' end and the left-hand direction of single or double-stranded nucleic acid sequences is referred to as the 5' direction.
  • the right-hand end or direction of a nucleic acid sequence is the 3' end or direction.
  • nucleotides that are upstream of a reference point in a nucleic acid may be designated by a negative number, while nucleotides that are downstream of a reference point may be designated by a positive number.
  • a reference point e.g., an exon-exon junction in mRNA
  • a nucleotide that is directly adjacent and upstream of the reference point is designated “minus one,” e.g., “-1,” while a nucleotide that is directly adjacent and downstream of the reference point is designated “plus one,” e.g., “+1.”
  • WSGR Docket No.47991-744.601 the ASOs are complementary to (and bind to) a targeted portion of a SCN1A NIE containing pre-mRNA that is downstream (in the 3' direction) of the 5' splice site (or 3' end of the NIE) of the included exon in a SCN1A NIE containing pre-mRNA (e.g., the direction designated by positive numbers relative to the 5' splice site).
  • the ASOs are complementary to a targeted portion of the SCN1A NIE containing pre-mRNA that is within the region about +1 to about +500 relative to the 5' splice site (or 3' end) of the included exon. In some embodiments, the ASOs may be complementary to a targeted portion of a SCN1A NIE containing pre-mRNA that is within the region between nucleotides +6 and +496 relative to the 5' splice site (or 3' end) of the included exon.
  • the ASOs are complementary to a targeted portion that is within the region about +1 to about +500, about +1 to about +490, about +1 to about +480, about +1 to about +470, about +1 to about +460, about +1 to about +450, about +1 to about +440, about +1 to about +430, about +1 to about +420, about +1 to about +410, about +1 to about +400, about +1 to about +390, about +1 to about +380, about +1 to about +370, about +1 to about +360, about +1 to about +350, about +1 to about +340, about +1 to about +330, about +1 to about +320, about +1 to about +310, about +1 to about +300, about +1 to about +290, about +1 to about +280, about +1 to about +270, about +1 to about +260, about +1 to about +250, about +1 to about +240, about +1 to about +230, about +1 to about +220,
  • the ASOs are complementary to a targeted portion that is within the region from about +1 to about +100, from about +100 to about +200, from about +200 to about +300, from about +300 to about +400, or from about +400 to about +500 relative to 5' splice site (or 3' end) of the included exon.
  • the ASOs are complementary to (and bind to) a targeted portion of a SCN1A NIE containing pre-mRNA that is upstream (in the 5' direction) of the 5' splice site (or 3' end) of the included exon in a SCN1A NIE containing pre-mRNA (e.g., the direction designated by negative numbers relative to the 5' splice site).
  • the ASOs are complementary to a targeted portion of the SCN1A NIE containing pre-mRNA that is within the region about -4 to about -270 relative to the 5' splice site (or 3' end) of the included exon.
  • the ASOs may be complementary to a targeted portion of a SCN1A NIE containing pre-mRNA that is within the region between nucleotides -1 and -264 relative to the 5' splice site (or 3' end) of the included exon.
  • the ASOs are complementary to a targeted portion that is within the region about -1 to about - 270, about -1 to about -260, about -1 to about -250, about -1 to about -240, about -1 to about -230, about -1 to about -220, about -1 to about -210, about -1 to about -200, about -1 to about -190, about -1 to about -180, about -1 to about -170, about -1 to about -160, about -1 to about -150, about -1 to about -140, about -1 to about -130, about -1 to about -120, about -1 to about -110, about -1 to about -100, about -1 to about -90, about -1 to about -80, about -1 to about -70, about -1 to about -60, about -1 to about -50, about -1 to WSGR Docket No.47991-744.601 about -40, about -1 to about a targeted portion
  • the ASOs are complementary to a targeted portion that is within the region from about -1 to about -50, from about -50 to about -100, from about -100 to about -150, from about -150 to about -200, or from about -200 to about -250 relative to 5' splice site (or 3' end) of the included exon.
  • the ASOs are complementary to a targeted region of a SCN1A NIE containing pre-mRNA that is upstream (in the 5' direction) of the 3' splice site (or 5' end) of the included exon in a SCN1A NIE containing pre-mRNA (e.g., in the direction designated by negative numbers).
  • the ASOs are complementary to a targeted portion of the SCN1A NIE containing pre-mRNA that is within the region about -1 to about -500 relative to the 3' splice site (or 5' end) of the included exon. In some embodiments, the ASOs are complementary to a targeted portion of the SCN1A NIE containing pre-mRNA that is within the region -1 to -496 relative to the 3' splice site of the included exon.
  • the ASOs are complementary to a targeted portion that is within the region about - 1 to about -500, about -1 to about -490, about -1 to about -480, about -1 to about -470, about -1 to about - 460, about -1 to about -450, about -1 to about -440, about -1 to about -430, about -1 to about -420, about -1 to about -410, about -1 to about -400, about -1 to about -390, about -1 to about -380, about -1 to about -370, about -1 to about -360, about -1 to about -350, about -1 to about -340, about -1 to about -330, about -1 to about -320, about -1 to about -310, about -1 to about -300, about -1 to about -290, about -1 to about -280, about -1 to about -270, about -1 to about -260, about -1 to about -250
  • the ASOs are complementary to a targeted portion that is within the region from about -1 to about -100, from about -100 to about -200, from about -200 to about -300, from about -300 to about - 400, or from about -400 to about -500 relative to 3' splice site of the included exon.
  • the ASOs are complementary to a targeted region of a SCN1A NIE containing pre-mRNA that is downstream (in the 3' direction) of the 3' splice site (5' end) of the included exon in a SCN1A NIE containing pre-mRNA (e.g., in the direction designated by positive numbers).
  • the ASOs are complementary to a targeted portion of the SCN1A NIE containing pre-mRNA that is within the region of about +1 to about +100 relative to the 3' splice site of the included exon. In some aspects, the ASOs are complementary to a targeted portion that is within the region about +1 to about +90, about +1 to about +80, about +1 to about +70, about +1 to about +60, about +1 to about +50, about +1 to about +40, about +1 to about +30, about +1 to about +20, or about +1 to about +10 relative to 3' splice site of the included exon.
  • the targeted portion of the SCN1A NIE containing pre-mRNA is within the region +100 relative to the 5' splice site (3' end) of the included exon to -100 relative to the 3' splice WSGR Docket No.47991-744.601 site (5' end) of the included exon.
  • the targeted portion of the SCN1A NIE containing pre-mRNA is within the NIE.
  • the targeted portion of the SCN1A NIE containing pre-mRNA comprises a pseudo-exon and intron boundary.
  • the ASOs may be of any length suitable for specific binding and effective enhancement of splicing.
  • the ASOs consist of 8 to 50 nucleobases.
  • the ASO may be 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 40, 45, or 50 nucleobases in length.
  • the ASOs consist of more than 50 nucleobases.
  • the ASO is from 8 to 50 nucleobases, 8 to 40 nucleobases, 8 to 35 nucleobases, 8 to 30 nucleobases, 8 to 25 nucleobases, 8 to 20 nucleobases, 8 to 15 nucleobases, 9 to 50 nucleobases, 9 to 40 nucleobases, 9 to 35 nucleobases, 9 to 30 nucleobases, 9 to 25 nucleobases, 9 to 20 nucleobases, 9 to 15 nucleobases, 10 to 50 nucleobases, 10 to 40 nucleobases, 10 to 35 nucleobases, 10 to 30 nucleobases, 10 to 25 nucleobases, 10 to 20 nucleobases, 10 to 15 nucleobases, 11 to 50 nucleobases, 11 to 40 nucleobases, 11 to 35 nucleobases, 11 to 30 nucleobases, 11 to 25 nucleobases, 11 to 20 nucleobases, 11 to
  • the ASOs are 18 nucleotides in length. In some embodiments, the ASOs are 15 nucleotides in length. In some embodiments, the ASOs are 25 nucleotides in length. [0595] In some embodiments, two or more ASOs with different chemistries but complementary to the same targeted portion of the NIE containing pre-mRNA are used. In some embodiments, two or more ASOs that are complementary to different targeted portions of the NIE containing pre-mRNA are used.
  • the antisense oligonucleotides or antisense oligomers of the invention are chemically linked to one or more moieties or conjugates, e.g., a targeting moiety or other conjugate that enhances the activity or cellular uptake of the oligonucleotide.
  • moieties include, but are not limited to, a lipid moiety, e.g., as a cholesterol moiety, a cholesteryl moiety, an aliphatic chain, e.g., dodecandiol or undecyl residues, a polyamine or a polyethylene glycol chain, or adamantane acetic acid.
  • the antisense oligonucleotide or antisense oligomer is conjugated with a moiety including, but not limited to, an abasic nucleotide, a polyether, a polyamine, a polyamide, a peptide, a carbohydrate, e.g., N-acetylgalactosamine (GalNAc), N-Ac-Glucosamine (GluNAc), or mannose (e.g., mannose-6-phosphate), a lipid, or a polyhydrocarbon compound.
  • a moiety including, but not limited to, an abasic nucleotide, a polyether, a polyamine, a polyamide, a peptide, a carbohydrate, e.g., N-acetylgalactosamine (GalNAc), N-Ac-Glucosamine (GluNAc), or mannose (e.g., mannose-6-phosphate), a lipid, or a polyhydr
  • Conjugates can be linked to one or more of any nucleotides comprising the antisense oligonucleotide or antisense oligomer at any of several positions on the sugar, base, or phosphate group, as understood in the art and described WSGR Docket No.47991-744.601 in the literature, e.g., using a linker.
  • Linkers can include a bivalent or trivalent branched linker.
  • the conjugate is attached to the 3' end of the antisense oligonucleotide or antisense oligomer. Methods of preparing oligonucleotide conjugates are described, e.g., in U.S. Pat. No.
  • the nucleic acid to be targeted by an ASO is a SCN1A NIE containing pre-mRNA expressed in a cell, such as a eukaryotic cell.
  • the term “cell” may refer to a population of cells.
  • the cell is in a subject.
  • the cell is isolated from a subject.
  • the cell is ex vivo.
  • the cell is a condition or disease-relevant cell or a cell line.
  • the cell is in vitro (e.g., in cell culture).
  • the ASO is the salt of a nucleotide. In some embodiments, the ASO is the salt of a nucleotide, fully phosphorothioate-linked oligonucleotide. In some embodiments, the ASO is the salt of a nucleotide in which the salt binds to the phosphate-link. In some embodiments, the ASO is the salt of a nucleotide, fully phosphorothioate-linked oligonucleotide in which the salt binds to the phosphate-link. In some embodiments, the ASO is the sodium salt of a nucleotide.
  • the ASO is the sodium salt of a nucleotide, fully phosphorothioate-linked oligonucleotide. In some embodiments, the ASO is the sodium salt of a nucleotide in which the sodium salt binds to the phosphate-link. In some embodiments, the ASO is the sodium salt of a nucleotide, fully phosphorothioate-linked oligonucleotide in which the sodium salt binds to the phosphate-link. In some embodiments, the ASO is the potassium salt of a nucleotide. In some embodiments, the ASO is the potassium salt of a nucleotide, fully phosphorothioate-linked oligonucleotide.
  • the ASO is the potassium salt of a nucleotide in which the potassium salt binds to the phosphate-link. In some embodiments, the ASO is the potassium salt of a nucleotide, fully phosphorothioate-linked oligonucleotide in which the potassium salt binds to the phosphate-link. [0599] In some embodiment, the ASO is the monosodium salt of a 2-nucleotide (2-mer). In some embodiment, the ASO is the disodium salt of a 3-nucleotide (3-mer). In some embodiment, the ASO is the trisodium salt of a 4-nucleotide (4-mer).
  • the ASO is the tetrasodium salt of a 5- nucleotide (5-mer). In some embodiment, the ASO is the pentasodium salt of a 6-nucleotide (6-mer). In some embodiment, the ASO is the hexasodium salt of a 7-nucleotide (7-mer). In some embodiment, the ASO is the heptasodium salt of a 8-nucleotide (8-mer). In some embodiment, the ASO is the octasodium salt of a 9-nucleotide (9-mer). In some embodiment, the ASO is the nonasodium salt of a 10-nucleotide (10-mer).
  • the ASO is the decasodium salt of a 11-nucleotide (11-mer). In some embodiment, the ASO is the undecasodium salt of a 12-nucleotide (12-mer). In some embodiment, the ASO is the dodecasodium salt of a 13-nucleotide (13-mer). In some embodiment, the ASO is the tridecasodium salt of a 14-nucleotide (14-mer). In some embodiment, the ASO is the tetradecasodium salt of a 15-nucleotide (15-mer). In some embodiment, the ASO is the pentadecasodium salt of a 16- nucleotide (16-mer).
  • the ASO is the hexadecasodium salt of a 17-nucleotide (17- WSGR Docket No.47991-744.601 mer). In some embodiment, the ASO is the heptadecasodium salt of a 18-nucleotide (18-mer). In some embodiment, the ASO is the octadecasodium salt of a 19-nucleotide (19-mer). In some embodiment, the ASO is the nonadecasodium salt of a 20-nucleotide (20-mer). In some embodiment, the ASO is the icosasodium salt of a 21-nucleotide (21-mer).
  • the ASO is the henicosasodium salt of a 22-nucleotide (22-mer). In some embodiment, the ASO is the docosasodium salt of a 23-nucleotide (23-mer). In some embodiment, the ASO is the tricosasodium salt of a 24-nucleotide (24-mer). In some embodiment, the ASO is the tetracosasodium salt of a 25-nucleotide (25-mer). In some embodiment, the ASO is the pentacosasodium salt of a 26-nucleotide (26-mer).
  • the ASO is the hexacosasodium salt of a 27-nucleotide (27-mer). In some embodiment, the ASO is the heptacosasodium salt of a 28-nucleotide (28-mer). In some embodiment, the ASO is the octacosasodium salt of a 29- nucleotide (29-mer). In some embodiment, the ASO is the nonacosasodium salt of a 30-nucleotide (30- mer). In some embodiment, the ASO is the triacontasodium salt of a 31-nucleotide (31-mer).
  • the ASO is the hentriacontasodium salt of a 32-nucleotide (32-mer). In some embodiment, the ASO is the dotriacontasodium salt of a 33-nucleotide (33-mer). In some embodiment, the ASO is the tritriacontasodium salt of a 34-nucleotide (34-mer). In some embodiment, the ASO is the tetratriacontasodium salt of a 35-nucleotide (35-mer). In some embodiment, the ASO is the pentatriacontasodium salt of a 36-nucleotide (36-mer).
  • the ASO is the hexatriacontasodium salt of a 37-nucleotide (37-mer). In some embodiment, the ASO is the heptatriacontasodium salt of a 38-nucleotide (38-mer). In some embodiment, the ASO is the octatriacontasodium salt of a 39-nucleotide (39-mer). In some embodiment, the ASO is the nonatriacontasodium salt of a 40-nucleotide (40-mer). In some embodiment, the ASO is the tetracontasodium salt of a 41-nucleotide (41-mer).
  • the ASO is the hentetracontasodium salt of a 42-nucleotide (42-mer). In some embodiment, the ASO is the dotetracontasodium salt of a 43-nucleotide (43-mer). In some embodiment, the ASO is the tritetracontasodium salt of a 44-nucleotide (44-mer). In some embodiment, the ASO is the tetratetracontasodium salt of a 45-nucleotide (45-mer). In some embodiment, the ASO is the pentatetracontasodium salt of a 46-nucleotide (46-mer).
  • the ASO is the hexatetracontasodium salt of a 47-nucleotide (47-mer). In some embodiment, the ASO is the heptatetracontasodium salt of a 48-nucleotide (48-mer). In some embodiment, the ASO is the octatetracontasodium salt of a 49-nucleotide (49-mer). In some embodiment, the ASO is the nonatetracontasodium salt of a 50-nucleotide (50-mer). In some embodiment, the ASO is the pentacontasodium salt of a 51-nucleotide (51-mer).
  • the ASO is the monosodium salt of a 2-nucleotide (2-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the disodium salt of a 3- nucleotide (3-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the trisodium salt of a 4-nucleotide (4-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tetrasodium salt of a 5-nucleotide (5-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the pentasodium salt of a 6-nucleotide (6-mer), fully WSGR Docket No.47991-744.601 phosphorothioate-linked oligonucleotide.
  • the ASO is the hexasodium salt of a 7- nucleotide (7-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the heptasodium salt of a 8-nucleotide (8-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the octasodium salt of a 9-nucleotide (9-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the nonasodium salt of a 10-nucleotide (10-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the decasodium salt of a 11- nucleotide (11-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the undecasodium salt of a 12-nucleotide (12-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the dodecasodium salt of a 13-nucleotide (13-mer), fully phosphorothioate- linked oligonucleotide.
  • the ASO is the tridecasodium salt of a 14-nucleotide (14- mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tetradecasodium salt of a 15-nucleotide (15-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the pentadecasodium salt of a 16-nucleotide (16-mer), fully phosphorothioate- linked oligonucleotide.
  • the ASO is the hexadecasodium salt of a 17-nucleotide (17- mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the heptadecasodium salt of an 18-nucleotide (18-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the octadecasodium salt of a 19-nucleotide (19-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the nonadecasodium salt of a 20-nucleotide (20-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the icosasodium salt of a 21-nucleotide (21-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the henicosasodium salt of a 22-nucleotide (22-mer), fully phosphorothioate- linked oligonucleotide.
  • the ASO is the docosasodium salt of a 23-nucleotide (23- mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tricosasodium salt of a 24-nucleotide (24-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tetracosasodium salt of a 25-nucleotide (25-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the pentacosasodium salt of a 26-nucleotide (26-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the hexacosasodium salt of a 27-nucleotide (27-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the heptacosasodium salt of a 28-nucleotide (28-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the octacosasodium salt of a 29-nucleotide (29-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the nonacosasodium salt of a 30-nucleotide (30-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the triacontasodium salt of a 31-nucleotide (31-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the hentriacontasodium salt of a 32-nucleotide (32- mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the dotriacontasodium salt of a 33-nucleotide (33-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tritriacontasodium salt of a 34-nucleotide (34-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tetratriacontasodium salt WSGR Docket No.47991-744.601 of a 35-nucleotide (35-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the pentatriacontasodium salt of a 36-nucleotide (36-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the hexatriacontasodium salt of a 37-nucleotide (37- mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the heptatriacontasodium salt of a 38-nucleotide (38-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the octatriacontasodium salt of a 39-nucleotide (39-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the nonatriacontasodium salt of a 40-nucleotide (40-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tetracontasodium salt of a 41-nucleotide (41-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the hentetracontasodium salt of a 42-nucleotide (42- mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the dotetracontasodium salt of a 43-nucleotide (43-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tritetracontasodium salt of a 44-nucleotide (44-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tetratetracontasodium salt of a 45-nucleotide (45-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the pentatetracontasodium salt of a 46-nucleotide (46-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the hexatetracontasodium salt of a 47-nucleotide (47- mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the heptatetracontasodium salt of a 48-nucleotide (48-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the octatetracontasodium salt of a 49-nucleotide (49-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the nonatetracontasodium salt of a 50-nucleotide (50-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the pentacontasodium salt of a 51-nucleotide (51-mer), fully phosphorothioate-linked oligonucleotide. [0601] In some embodiment, the ASO is the monopotassium salt of a 2-nucleotide (2-mer). In some embodiment, the ASO is the dipotassium salt of a 3-nucleotide (3-mer).
  • the ASO is the tripotassium salt of a 4-nucleotide (4-mer). In some embodiment, the ASO is the tetrapotassium salt of a 5-nucleotide (5-mer). In some embodiment, the ASO is the pentapotassium salt of a 6-nucleotide (6- mer). In some embodiment, the ASO is the hexapotassium salt of a 7-nucleotide (7-mer). In some embodiment, the ASO is the heptapotassium salt of an 8-nucleotide (8-mer). In some embodiment, the ASO is the octapotassium salt of a 9-nucleotide (9-mer).
  • the ASO is the nonapotassium salt of a 10-nucleotide (10-mer). In some embodiment, the ASO is the decapotassium salt of a 11-nucleotide (11-mer). In some embodiment, the ASO is the undecapotassium salt of a 12- nucleotide (12-mer). In some embodiment, the ASO is the dodecapotassium salt of a 13-nucleotide (13- mer). In some embodiment, the ASO is the tridecapotassium salt of a 14-nucleotide (14-mer). In some embodiment, the ASO is the tetradecapotassium salt of a 15-nucleotide (15-mer).
  • the ASO is the pentadecapotassium salt of a 16-nucleotide (16-mer). In some embodiment, the ASO is the hexadecapotassium salt of a 17-nucleotide (17-mer). In some embodiment, the ASO is the WSGR Docket No.47991-744.601 heptadecapotassium salt of a 18-nucleotide (18-mer). In some embodiment, the ASO is the octadecapotassium salt of a 19-nucleotide (19-mer). In some embodiment, the ASO is the nonadecapotassium salt of a 20-nucleotide (20-mer).
  • the ASO is the icosapotassium salt of a 21-nucleotide (21-mer). In some embodiment, the ASO is the henicosapotassium salt of a 22-nucleotide (22-mer). In some embodiment, the ASO is the docosapotassium salt of a 23- nucleotide (23-mer). In some embodiment, the ASO is the tricosapotassium salt of a 24-nucleotide (24- mer). In some embodiment, the ASO is the tetracosapotassium salt of a 25-nucleotide (25-mer).
  • the ASO is the pentacosapotassium salt of a 26-nucleotide (26-mer). In some embodiment, the ASO is the hexacosapotassium salt of a 27-nucleotide (27-mer). In some embodiment, the ASO is the heptacosapotassium salt of a 28-nucleotide (28-mer). In some embodiment, the ASO is the octacosapotassium salt of a 29-nucleotide (29-mer). In some embodiment, the ASO is the nonacosapotassium salt of a 30-nucleotide (30-mer).
  • the ASO is the triacontapotassium salt of a 31-nucleotide (31-mer). In some embodiment, the ASO is the hentriacontapotassium salt of a 32-nucleotide (32-mer). In some embodiment, the ASO is the dotriacontapotassium salt of a 33-nucleotide (33-mer). In some embodiment, the ASO is the tritriacontapotassium salt of a 34-nucleotide (34-mer). In some embodiment, the ASO is the tetratriacontapotassium salt of a 35-nucleotide (35-mer).
  • the ASO is the pentatriacontapotassium salt of a 36-nucleotide (36-mer). In some embodiment, the ASO is the hexatriacontapotassium salt of a 37-nucleotide (37-mer). In some embodiment, the ASO is the heptatriacontapotassium salt of a 38-nucleotide (38-mer). In some embodiment, the ASO is the octatriacontapotassium salt of a 39-nucleotide (39-mer). In some embodiment, the ASO is the nonatriacontapotassium salt of a 40-nucleotide (40-mer).
  • the ASO is the tetracontapotassium salt of a 41-nucleotide (41-mer). In some embodiment, the ASO is the hentetracontapotassium salt of a 42-nucleotide (42-mer). In some embodiment, the ASO is the dotetracontapotassium salt of a 43-nucleotide (43-mer). In some embodiment, the ASO is the tritetracontapotassium salt of a 44-nucleotide (44-mer). In some embodiment, the ASO is the tetratetracontapotassium salt of a 45-nucleotide (45-mer).
  • the ASO is the pentatetracontapotassium salt of a 46-nucleotide (46-mer). In some embodiment, the ASO is the hexatetracontapotassium salt of a 47-nucleotide (47-mer). In some embodiment, the ASO is the heptatetracontapotassium salt of a 48-nucleotide (48-mer). In some embodiment, the ASO is the octatetracontapotassium salt of a 49-nucleotide (49-mer). In some embodiment, the ASO is the nonatetracontapotassium salt of a 50-nucleotide (50-mer).
  • the ASO is the pentacontapotassium salt of a 51-nucleotide (51-mer).
  • the ASO is the monopotassium salt of a 2-nucleotide (2-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the dipotassium salt of a 3- nucleotide (3-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tripotassium salt of a 4-nucleotide (4-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tetrapotassium salt of a 5-nucleotide (5-mer), fully phosphorothioate-linked WSGR Docket No.47991-744.601 oligonucleotide.
  • the ASO is the pentapotassium salt of a 6-nucleotide (6-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the hexapotassium salt of a 7-nucleotide (7-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the heptapotassium salt of an 8-nucleotide (8-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the octapotassium salt of a 9-nucleotide (9-mer), fully phosphorothioate- linked oligonucleotide. In some embodiment, the ASO is the nonapotassium salt of a 10-nucleotide (10- mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the decapotassium salt of a 11-nucleotide (11-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the undecapotassium salt of a 12-nucleotide (12-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the dodecapotassium salt of a 13-nucleotide (13-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tridecapotassium salt of a 14-nucleotide (14-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tetradecapotassium salt of a 15-nucleotide (15-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the pentadecapotassium salt of a 16-nucleotide (16- mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the hexadecapotassium salt of a 17-nucleotide (17-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the heptadecapotassium salt of a 18-nucleotide (18-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the octadecapotassium salt of a 19-nucleotide (19-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the nonadecapotassium salt of a 20-nucleotide (20-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the icosapotassium salt of a 21-nucleotide (21-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the henicosapotassium salt of a 22-nucleotide (22-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the docosapotassium salt of a 23-nucleotide (23-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tricosapotassium salt of a 24-nucleotide (24-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tetracosapotassium salt of a 25-nucleotide (25-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the pentacosapotassium salt of a 26-nucleotide (26-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the hexacosapotassium salt of a 27-nucleotide (27- mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the heptacosapotassium salt of a 28-nucleotide (28-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the octacosapotassium salt of a 29-nucleotide (29-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the nonacosapotassium salt of a 30-nucleotide (30-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the triacontapotassium salt of a 31-nucleotide (31-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the hentriacontapotassium salt of a 32-nucleotide (32-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the dotriacontapotassium salt of a 33-nucleotide (33-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the WSGR Docket No.47991-744.601 ASO is the tritriacontapotassium salt of a 34-nucleotide (34-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tetratriacontapotassium salt of a 35-nucleotide (35- mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the pentatriacontapotassium salt of a 36-nucleotide (36-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the hexatriacontapotassium salt of a 37-nucleotide (37-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the heptatriacontapotassium salt of a 38-nucleotide (38-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the octatriacontapotassium salt of a 39-nucleotide (39-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the nonatriacontapotassium salt of a 40-nucleotide (40-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tetracontapotassium salt of a 41-nucleotide (41-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the hentetracontapotassium salt of a 42-nucleotide (42-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the dotetracontapotassium salt of a 43-nucleotide (43-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the tritetracontapotassium salt of a 44-nucleotide (44-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the tetratetracontapotassium salt of a 45-nucleotide (45-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the pentatetracontapotassium salt of a 46-nucleotide (46-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the hexatetracontapotassium salt of a 47-nucleotide (47-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the heptatetracontapotassium salt of a 48-nucleotide (48-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the octatetracontapotassium salt of a 49-nucleotide (49-mer), fully phosphorothioate-linked oligonucleotide.
  • the ASO is the nonatetracontapotassium salt of a 50-nucleotide (50-mer), fully phosphorothioate-linked oligonucleotide. In some embodiment, the ASO is the pentacontapotassium salt of a 51-nucleotide (51-mer), fully phosphorothioate-linked oligonucleotide. [0603] In some embodiments, the ASO as described herein is a compound having the structure depicted in formula (I) (free acid):
  • the ASO as described herein is a compound having the structure depicted in formula (II) (sodium salt): (II).
  • an ASO described herein is all-P-ambo-2'-O-(2-methoxyethyl)-P-thioadenylyl- (3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-5-methyl-P-thiouridylyl- (3' ⁇ 5')-2'-O-(2-methoxyethyl)-5-methyl-P-thiouridylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl- (3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl- (3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyeth
  • an ASO described herein is a sodium salt of all-P-ambo-2'-O-(2-methoxyethyl)- P-thioadenylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-5-methyl- P-thiouridylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-5-methyl-P-thiouridylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)- P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-O-(2-methoxyethyl)-P-thioguanylyl-(3' ⁇ 5')-2'-
  • SCN1A sodium channel, voltage-gated, type I, alpha subunit
  • SCN1A mutations in DS are spread across the entire protein. More than 100 novel mutations have been identified throughout the gene with the more debilitating arising de novo. These comprise of truncations (47%), missense (43%), deletions (3%), and splice site mutations (7%). The percentage of subjects carrying SCN1A mutations varies between 33 and 100%. The majority of mutations are novel changes (88%).
  • the methods described herein are used to modulate, e.g., increase or decrease, the production of a functional NaV1.1 protein.
  • the term “functional” refers to the amount of activity or function of a NaV1.1 protein that is necessary to eliminate any one or more symptoms of a treated condition, e.g., Dravet syndrome; Epilepsy, generalized, with febrile seizures plus, type 2; Febrile seizures, familial, 3A; Autism; Epileptic encephalopathy, early infantile, 13; Sick sinus syndrome 1; Alzheimer's disease; or SUDEP.
  • the methods are used to increase the production of a partially functional NaV1.1 protein.
  • partially functional refers to any amount of activity or function of the NaV1.1 protein that is less than the amount of activity or function that is necessary to eliminate or prevent any one or more symptoms of a disease or condition.
  • a partially functional protein or RNA will have at least 10%, at least 20%, at least WSGR Docket No.47991-744.601 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% less activity relative to the fully functional protein or RNA.
  • the method is a method of increasing the expression of the NaV1.1 protein by cells of a subject having a NIE containing pre-mRNA encoding the NaV1.1 protein, wherein the subject has Dravet syndrome caused by a deficient amount of activity of NaV1.1 protein, and wherein the deficient amount of the NaV1.1 protein is caused by haploinsufficiency of the NaV1.1 protein.
  • the subject has a first allele encoding a functional NaV1.1 protein, and a second allele from which the NaV1.1 protein is not produced.
  • the subject has a first allele encoding a functional NaV1.1 protein, and a second allele encoding a nonfunctional NaV1.1 protein.
  • the subject has a first allele encoding a functional NaV1.1 protein, and a second allele encoding a partially functional NaV1.1 protein.
  • the subject expresses a partially functional Na V 1.1 protein from one allele, wherein the partially functional Na V 1.1 protein is caused by a frameshift mutation, a non-sense mutation, a missense mutation, or a partial gene deletion.
  • the subject expresses a nonfunctional Na V 1.1 protein from one allele, wherein the nonfunctional Na V 1.1 protein is caused by a frameshift mutation, a nonsense mutation, a missense mutation, a partial gene deletion, in one allele.
  • the subject has a SCN1A whole gene deletion, in one allele.
  • the antisense oligomer binds to a targeted portion of the NIE containing pre-mRNA transcribed from the second allele, thereby inducing exon skipping of the pseudo-exon from the pre-mRNA, and causing an increase in the level of mature mRNA encoding functional Na V 1.1 protein, and an increase in the expression of the Na V 1.1 protein in the cells of the subject.
  • a subject can have a mutation in SCN1A. Mutations in SCN1A can be spread throughout said gene. Na V 1.1 protein can consist of four domains.
  • Said SCN1A domains can have transmembrane segments. Mutations in said Na V 1.1 protein may arise throughout said protein. Said Na V 1.1 protein may consist of at least two isoforms. Mutations in SCN1A may comprise of R931C, R946C, M934I, R1648C, or R1648H. In some cases, mutations may be observed in a C-terminus of a Na V 1.1 protein. Mutations in a Na V 1.1 protein may also be found in loops between segments 5 and 6 of the first three domains of said Na V 1.1 protein. In some cases, mutations may be observed in an N- terminus of a Na V 1.1 protein.
  • Exemplary mutations within SCN1A include, but are not limited to, R222X, R712X, I227S, R1892X, W952X, R1245X, R1407X, W1434R, c.4338+1G>A, 51516X, L1670fsX1678, or K1846fsX1856. Mutations that can be targeted with the present invention may also encode a pore of an ion channel. [0611] In some embodiments, the methods and compositions described herein can be used to treat DS. In other embodiments, the methods and compositions described herein can be used to treat severe myclonic epilepsy of infancy (SMEI).
  • SMEI severe myclonic epilepsy of infancy
  • the methods and compositions described herein can be used to treat borderline Dravet syndrome; Epilepsy, generalized, with febrile seizures plus, type 2; Febrile seizures, familial, 3A; Migraine, familial hemiplegic, 3; Autism; Epileptic encephalopathy, early infantile, 13; Sick sinus syndrome 1; Alzheimer’s disease or SUDEP.
  • WSGR Docket No.47991-744.601 the method is a method of using an ASO to increase the expression of a protein or functional RNA.
  • an ASO is used to increase the expression of NaV1.1 protein in cells of a subject having a NIE containing pre-mRNA encoding NaV1.1 protein, wherein the subject has a deficiency, e.g., Dravet Syndrome (DS) (also known as SMEI); severe myoclonic epilepsy of infancy (SMEI)-borderland (SMEB); Febrile seizure (FS); epilepsy, generalized, with febrile seizures plus (GEFS+); epileptic encephalopathy, early infantile, 13; cryptogenic generalized epilepsy; cryptogenic focal epilepsy; myoclonic-astatic epilepsy; Lennox-Gastaut syndrome; West syndrome; idiopathic spasms; early myoclonic encephalopathy; progressive myoclonic epilepsy; alternating hemiplegia of childhood; unclassified epileptic encephalopathy; sudden unexpected death in epilepsy (SUDEP); sick sinus syndrome 1; early infantile SCN1
  • DS
  • an ASO is used to increase the expression of Na V 1.1 protein in cells of a subject, wherein the subject has a deficiency, e.g., Epileptic encephalopathy, early infantile, 13; in the amount or function of a SCN8A protein.
  • an ASO is used to increase the expression of Na V 1.1 protein in cells of a subject, wherein the subject has a deficiency, e.g., Sick sinus syndrome 1; in the amount or function of a SCN5A protein.
  • a subject having any SCN1A mutation known in the art and described in the literature e.g., by Hamdan, et al., 2009, N. Engl. Med.
  • the mutation is within any SCN1A intron or exon.
  • the NIE containing pre-mRNA transcript that encodes the protein that is causative of the disease or condition is targeted by the ASOs described herein.
  • a NIE containing pre-mRNA transcript that encodes a protein that is not causative of the disease is targeted by the ASOs.
  • a disease that is the result of a mutation or deficiency of a first protein in a particular pathway may be ameliorated by targeting a NIE containing pre-mRNA that encodes a second protein, thereby increasing production of the second protein.
  • the function of the second protein is able to compensate for the mutation or deficiency of the first protein (which is causative of the disease or condition).
  • the subject has: (a) a first mutant allele from which (i) the NaV1.1 protein is produced at a reduced level compared to production from a wild-type allele, (ii) the NaV1.1 protein is produced in a form having reduced function compared to an equivalent wild-type protein, or (iii) the NaV1.1 protein or functional RNA is not produced; and (b) a second mutant allele from which WSGR Docket No.47991-744.601 (i) the NaV1.1 protein is produced at a reduced level compared to production from a wild-type allele, (ii) the NaV1.1 protein is produced in a form having reduced function compared to an equivalent wild-type protein, or (iii) the NaV1.1 protein is not produced, and wherein the NIE containing pre-mRNA is transcribed from the first allele and/or the second allele.
  • the ASO binds to a targeted portion of the NIE containing pre-mRNA transcribed from the first allele or the second allele, thereby inducing exon skipping of the pseudo-exon from the NIE containing pre-mRNA, and causing an increase in the level of mRNA encoding NaV1.1 protein and an increase in the expression of the target protein or functional RNA in the cells of the subject.
  • the target protein or functional RNA having an increase in expression level resulting from the exon skipping of the pseudo-exon from the NIE containing pre-mRNA is either in a form having reduced function compared to the equivalent wild-type protein (partially-functional), or having full function compared to the equivalent wild-type protein (fully-functional).
  • the level of mRNA encoding Na V 1.1 protein is increased 1.1 to 10-fold, when compared to the amount of mRNA encoding Na V 1.1 protein that is produced in a control cell, e.g., one that is not treated with the antisense oligomer or one that is treated with an antisense oligomer that does not bind to the targeted portion of the SCN1A NIE containing pre-mRNA.
  • a subject treated using the methods of the present disclosure expresses a mutant Na V 1.1 protein from one allele, wherein the mutant Na V 1.1 protein is caused by a frameshift mutation, a nonsense mutation, a missense mutation, or a partial gene deletion, and wherein the mutant Na V 1.1 protein causes an elevated activity level of Na V 1.1.
  • a subject treated using the methods of the present disclosure expresses an elevated amount of Na V 1.1 protein from one allele due to a frameshift mutation, a nonsense mutation, a missense mutation, or a partial gene deletion.
  • a subject treated using the methods of the present disclosure expresses a partially functional Na V 1.1 protein from one allele, wherein the partially functional Na V 1.1 protein is caused by a frameshift mutation, a nonsense mutation, a missense mutation, or a partial gene deletion.
  • a subject treated using the methods of the disclosure expresses a nonfunctional Na V 1.1 protein from one allele, wherein the nonfunctional Na V 1.1 protein is caused by a frameshift mutation, a nonsense mutation, a missense mutation, a partial gene deletion, in one allele.
  • a subject treated using the methods of the disclosure has a SCN1A whole gene deletion, in one allele.
  • the method is a method of decreasing the expression of the NaV1.1 protein by cells of a subject having a NIE containing pre-mRNA encoding the NaV1.1 protein, and wherein the subject has a gain-of-function mutation in NaV1.1.
  • the subject has an allele from which the NaV1.1 protein is produced in an elevated amount or an allele encoding a mutant SCN1A that induces increased activity of NaV1.1 in the cell.
  • the increased activity of NaV1.1 is characterized by a prolonged or near persistent sodium current mediated by the mutant NaV1.1 channel, a WSGR Docket No.47991-744.601 slowing of fast inactivation, a positive shift in steady-state inactivation, higher channel availability during repetitive stimulation, increased non-inactivated depolarization-induced persistent sodium currents, delayed entry into inactivation, accelerated recovery from fast inactivation, and/or rescue of folding defects by incubation at lower temperature or co-expression of interacting proteins.
  • Splicing of the identified SCN1A NIE pre-mRNA species to produce functional mature SCN1A mRNA can be induced using a therapeutic agent such as an ASO that stimulates exon skipping of an NIE. Induction of exon skipping can result in inhibition of an NMD pathway.
  • the resulting mature SCN1A mRNA can be translated normally without activating NMD pathway, thereby increasing the amount of NaV1.1 protein in the patient's cells and alleviating symptoms of a condition associated with SCN1A deficiency, such as Dravet Syndrome (DS); Epilepsy, generalized, with febrile seizures plus, type 2; Febrile seizures, familial, 3A; Autism; Epileptic encephalopathy, early infantile, 13; Sick sinus syndrome 1; Alzheimer's disease; or SUDEP.
  • DS Dravet Syndrome
  • Epilepsy generalized, with febrile seizures plus, type 2; Febrile seizures, familial, 3A; Autism; Epileptic encephalopathy, early infantile, 13; Sick sinus syndrome 1; Alzheimer's disease; or SUDEP.
  • SCN1A pre-mRNA transcripts to modulate, e.g., enhance or inhibit, splicing or protein expression level.
  • the therapeutic agent can be a small molecule, polynucleotide, or polypeptide.
  • the therapeutic agent is an ASO.
  • Various regions or sequences on the SCN1A pre-mRNA can be targeted by a therapeutic agent, such as an ASO.
  • the ASO targets a SCN1A pre-mRNA transcript containing an NIE.
  • the ASO targets a sequence within an NIE of a SCN1A pre-mRNA transcript.
  • the ASO targets a sequence upstream (or 5') from the 5' end of an NIE (3'ss) of a SCN1A pre-mRNA transcript.
  • the ASO targets a sequence downstream (or 3') from the 3' end of an NIE (5'ss) of a SCN1A pre-mRNA transcript. In some embodiments, the ASO targets a sequence that is within an intron flanking on the 5' end of the NIE of a SCN1A pre-mRNA transcript. In some embodiments, the ASO targets a sequence that is within an intron flanking the 3' end of the NIE of a SCN1A pre-mRNA transcript. In some embodiments, the ASO targets a sequence comprising an NIE-intron boundary of a SCN1A pre-mRNA transcript. An NIE-intron boundary can refer to the junction of an intron sequence and an NIE region.
  • the intron sequence can flank the 5' end of the NIE, or the 3' end of the NIE.
  • the ASO targets a sequence within an exon of a SCN1A pre-mRNA transcript.
  • the ASO targets a sequence within an intron of a SCN1A pre-mRNA transcript.
  • the ASO targets a sequence comprising both a portion of an intron and a portion of an exon.
  • a therapeutic agent described herein modulates binding of a factor involved in splicing of the pre-mRNA containing an NMD exon.
  • a therapeutic agent described herein interferes with binding of a factor involved in splicing of the pre-mRNA containing an NMD exon. In some embodiments, a therapeutic agent described herein prevents binding of a factor involved in splicing of the pre-mRNA containing an NMD exon. In some embodiments, a therapeutic agent targets a targeted portion located in an intronic region between two canonical exonic regions of the pre-mRNA containing an NMD exon and encoding NaV1.1, and wherein the intronic WSGR Docket No.47991-744.601 region contains the NMD exon. In some embodiments, a therapeutic agent targets a targeted portion at least partially overlaps with the NMD exon.
  • a therapeutic agent targets a targeted portion that is at least partially overlaps with an intron upstream of the NMD exon. In some embodiments, a therapeutic agent targets a targeted portion within the NMD exon. [0623] In some embodiments, a therapeutic agent targets a targeted portion comprising at least about 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or more consecutive nucleotides of the NMD exon. In some embodiments, a therapeutic agent targets a targeted portion comprising at most about 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or more consecutive nucleotides of the NMD exon.
  • a therapeutic agent targets a targeted portion comprising about 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or more consecutive nucleotides of the NMD exon. [0624] In some embodiments, a therapeutic agent targets a targeted portion proximal to the NMD exon. [0625] In some embodiments, the ASO targets a sequence from about 4 to about 300 nucleotides upstream (or 5') from the 5' end of the NIE.
  • the ASO targets a sequence from about 1 to about 20 nucleotides, about 20 to about 50 nucleotides, about 50 to about 100 nucleotides, about 100 to about 150 nucleotides, about 150 to about 200 nucleotides, about 200 to about 250 nucleotides, about 250 to about 300, about 250 to about 300 nucleotides, about 350 to about 400 nucleotides, about 450 to about 500 nucleotides, about 550 to about 600 nucleotides, about 650 to about 700 nucleotides, about 750 to about 800 nucleotides, about 850 to about 900 nucleotides, about 950 to about 1000 nucleotides, about 1050 to about 1100 nucleotides, about 1150 to about 1200 nucleotides, about 1250 to about 1300 nucleotides, about 1350 to about 1400 nucleotides, or about 1450 to about 1500 nucleotides upstream (or 5') from the 5'
  • the ASO may target a sequence more than 300 nucleotides upstream from the 5' end of the NIE. In some embodiments, the ASO targets a sequence from about 4 to about 300 nucleotides downstream (or 3') from the 3' end of the NIE.
  • the ASO targets a sequence about 1 to about 20 nucleotides, about 20 to about 50 nucleotides, about 50 to about 100 nucleotides, about 100 to about 150 nucleotides, about 150 to about 200 nucleotides, about 200 to about 250 nucleotides, about 250 to about 300 nucleotides, about 350 to about 400 nucleotides, about 450 to about 500 nucleotides, about 550 to about 600 nucleotides, about 650 to about 700 nucleotides, about 750 to about 800 nucleotides, about 850 to about 900 nucleotides, about 950 to about 1000 nucleotides, about 1050 to about 1100 nucleotides, about 1150 to about 1200 nucleotides, about 1250 to about 1300 nucleotides, about 1350 to about 1400 nucleotides, or about 1450 to about 1500 nucleotides downstream from the 3' end of the NIE.
  • the ASO targets a sequence more than 300 nucleotides downstream from the 3' end of the NIE. [0626] In some embodiments, the ASO targets a sequence from about 4 to about 300 nucleotides upstream (or 5') from the 5' end of the NIE.
  • the ASO targets a sequence at least about 1 nucleotide, at least about 10 nucleotides, at least about 20 nucleotides, at least about 50 nucleotides, at least about 80 nucleotides, at least about 85 nucleotides, at least about 90 nucleotides, at least about 95 nucleotides, at least about 96 nucleotides, at least about 97 nucleotides, at least about 98 WSGR Docket No.47991-744.601 nucleotides, at least about 99 nucleotides, at least about 100 nucleotides, at least about 101 nucleotides, at least about 102 nucleotides, at least about 103 nucleotides, at least about 104 nucleotides, at least about 105 nucleotides, at least about 110 nucleotides, at least about 120 nucleotides, at least about 150 nucleotides, at least about 200 nucleotides, at
  • the ASO targets a sequence about 4 to about 300 nucleotides downstream (or 3') from the 3' end of the NIE. In some embodiments, the ASO targets a sequence at least about 1 nucleotide, at least about 10 nucleotides, at least about 20 nucleotides, at least about 50 nucleotides, at least about 80 nucleotides, at least about 85 nucleotides, at least about 90 nucleotides, at least about 95 nucleotides, at least about 96 nucleotides, at least about 97 nucleotides, at least about 98 nucleotides, at least about 99 nucleotides, at least about 100 nucleotides, at least about 101 nucleotides, at least about 102 nucleotides, at least about 103 nucleotides, at least about 104 nucleotides, at least about 105 nucleotides, at least about 110 nucleotides, at least about 120 nucleot
  • the ASO targets a sequence more than 300 nucleotides downstream from the 3' end of the NIE. [0627] In some embodiments, the ASO targets a sequence from about 4 to about 300 nucleotides upstream (or 5') from the 5' end of the NIE.
  • the ASO targets a sequence at most about 10 nucleotides, at most about 20 nucleotides, at most about 50 nucleotides, at most about 80 nucleotides, at most about 85 nucleotides, at most about 90 nucleotides, at most about 95 nucleotides, at most about 96 nucleotides, at most about 97 nucleotides, at most about 98 nucleotides, at most about 99 nucleotides, at most about 100 nucleotides, at most about 101 nucleotides, at most about 102 nucleotides, at most about 103 nucleotides, at most about 104 nucleotides, at most about 105 nucleotides, at most about 110 nucleotides, at most about 120 nucleotides, at most about 150 nucleotides, at most about 200 nucleotides, at most about 300 nucleotides, at most about 400 nucleotides, at most about 500 nucle
  • the ASO targets a sequence about 4 to about 300 nucleotides downstream (or 3') from the 3' end of the NIE. In some embodiments, the ASO targets a sequence at most about 10 nucleotides, at most about 20 nucleotides, at most about 50 nucleotides, at most about 80 nucleotides, at most about 85 nucleotides, at most about 90 nucleotides, at most about 95 nucleotides, at most about 96 nucleotides, at most about 97 nucleotides, at most about 98 nucleotides, at most about 99 nucleotides, at most about 100 nucleotides, at most about 101 nucleotides, at most about 102 nucleotides, at most about 103 nucleotides, at most about 104 WSGR Docket No.47991-744.601 nucleotides, at most about 105 nucleotides, at most about 110 nucleotides, at most about
  • the ASO targets a sequence more than 300 nucleotides downstream from the 3' end of the NIE.
  • the NIE as described herein is located between GRCh37/hg19: chr2:166,863,740 and GRCh37/hg19: chr2:166,863,803, as depicted in FIG.2.
  • the 5' end of the NIE is located at GRCh37/hg19: chr2:166,863,803.
  • the 3' end of the NIE is located at GRCh37/hg19: chr2:166,863,740.
  • the ASO targets a sequence from about 4 to about 300 nucleotides upstream (or 5') from genomic site GRCh37/hg19: chr2:166,863,803. In some embodiments, the ASO targets a sequence about 1 to about 20 nucleotides, about 20 to about 50 nucleotides, about 50 to about 100 nucleotides, about 100 to about 150 nucleotides, about 150 to about 200 nucleotides, about 200 to about 250 nucleotides, about 250 to about 300, about 250 to about 300 nucleotides, about 350 to about 400 nucleotides, about 450 to about 500 nucleotides, about 550 to about 600 nucleotides, about 650 to about 700 nucleotides, about 750 to about 800 nucleotides, about 850 to about 900 nucleotides, about 950 to about 1000 nucleotides, about 1050 to about 1100 nucleotides, about 1150 to about 1
  • the ASO may target a sequence more than 300 nucleotides upstream from genomic site GRCh37/hg19: chr2:166,863,803. In some embodiments, the ASO targets a sequence from about 4 to about 300 nucleotides downstream (or 3') from GRCh37/hg19: chr2:166,863,740.
  • the ASO targets a sequence about 1 to about 20 nucleotides, about 20 to about 50 nucleotides, about 50 to about 100 nucleotides, about 100 to about 150 nucleotides, about 150 to about 200 nucleotides, about 200 to about 250 nucleotides, about 250 to about 300 nucleotides, about 350 to about 400 nucleotides, about 450 to about 500 nucleotides, about 550 to about 600 nucleotides, about 650 to about 700 nucleotides, about 750 to about 800 nucleotides, about 850 to about 900 nucleotides, about 950 to about 1000 nucleotides, about 1050 to about 1100 nucleotides, about 1150 to about 1200 nucleotides, about 1250 to about 1300 nucleotides, about 1350 to about 1400 nucleotides, or about 1450 to about 1500 nucleotides downstream from GRCh37/hg19: chr2:166,
  • the ASO targets a sequence more than 300 nucleotides downstream from GRCh37/hg19: chr2: 166,863,740. [0630] In some embodiments, the ASO targets a sequence from about 4 to about 300 nucleotides upstream (or 5') from genomic site GRCh37/hg19: chr2:166,863,803.
  • the ASO targets a sequence at least about 1 nucleotide, at least about 10 nucleotides, at least about 20 nucleotides, at least about 50 nucleotides, at least about 80 nucleotides, at least about 85 nucleotides, at least about 90 WSGR Docket No.47991-744.601 nucleotides, at least about 95 nucleotides, at least about 96 nucleotides, at least about 97 nucleotides, at least about 98 nucleotides, at least about 99 nucleotides, at least about 100 nucleotides, at least about 101 nucleotides, at least about 102 nucleotides, at least about 103 nucleotides, at least about 104 nucleotides, at least about 105 nucleotides, at least about 110 nucleotides, at least about 120 nucleotides, at least about 150 nucleotides, at least about 200 nucleotides, at
  • the ASO targets a sequence from about 4 to about 300 nucleotides downstream (or 3') from GRCh37/hg19: chr2:166,863,740. In some embodiments, the ASO targets a sequence at least about 1 nucleotide, at least about 10 nucleotides, at least about 20 nucleotides, at least about 50 nucleotides, at least about 80 nucleotides, at least about 85 nucleotides, at least about 90 nucleotides, at least about 95 nucleotides, at least about 96 nucleotides, at least about 97 nucleotides, at least about 98 nucleotides, at least about 99 nucleotides, at least about 100 nucleotides, at least about 101 nucleotides, at least about 102 nucleotides, at least about 103 nucleotides, at least about 104 nucleotides, at least about 105 nucleotides, at least about 110 nucleotides
  • the ASO targets a sequence more than 300 nucleotides downstream from GRCh37/hg19: chr2:166,863,740. [0631] In some embodiments, the ASO targets a sequence from about 4 to about 300 nucleotides upstream (or 5') from genomic site GRCh37/hg19: chr2:166,863,803.
  • the ASO targets a sequence at most about 10 nucleotides, at most about 20 nucleotides, at most about 50 nucleotides, at most about 80 nucleotides, at most about 85 nucleotides, at most about 90 nucleotides, at most about 95 nucleotides, at most about 96 nucleotides, at most about 97 nucleotides, at most about 98 nucleotides, at most about 99 nucleotides, at most about 100 nucleotides, at most about 101 nucleotides, at most about 102 nucleotides, at most about 103 nucleotides, at most about 104 nucleotides, at most about 105 nucleotides, at most about 110 nucleotides, at most about 120 nucleotides, at most about 150 nucleotides, at most about 200 nucleotides, at most about 300 nucleotides, at most about 400 nucleotides, at most about 500 nucle
  • the ASO targets a sequence from about 4 to about 300 nucleotides downstream (or 3') from GRCh37/hg19: chr2:166,863,740. In some embodiments, the ASO targets a sequence at most about 10 nucleotides, at most about 20 nucleotides, at most about 50 nucleotides, at most about 80 nucleotides, at most about 85 nucleotides, at most about 90 nucleotides, at WSGR Docket No.47991-744.601 most about 95 nucleotides, at most about 96 nucleotides, at most about 97 nucleotides, at most about 98 nucleotides, at most about 99 nucleotides, at most about 100 nucleotides, at most about 101 nucleotides, at most about 102 nucleotides, at most about 103 nucleotides, at most about 104 nucleotides, at most about 105 nucleotides,
  • the ASO targets a sequence more than 300 nucleotides downstream from GRCh37/hg19: chr2:166,863,740.
  • the SCN1A gene (SEQ ID NO.1) was analyzed for NIE and inclusion of a portion of intron 20 (SEQ ID NO.4) (this portion is referred as exon 20x throughout the present disclosure) was observed.
  • the ASOs disclosed herein target a NIE containing pre-mRNA (SEQ ID NO.2) transcribed from a SCN1A genomic sequence.
  • the ASO targets a NIE containing pre-mRNA transcript from a SCN1A genomic sequence comprising a portion of intron 20.
  • the ASO targets a NIE containing pre-mRNA transcript from a SCN1A genomic sequence comprising exon 20x (SEQ ID NO.6). In some embodiments, the ASO targets a NIE containing pre- mRNA transcript of SEQ ID NO.2 or 12. In some embodiments, the ASO targets a NIE containing pre- mRNA transcript of SEQ ID NO.2 or 12 comprising an NIE. In some embodiments, the ASO targets a NIE containing pre-mRNA transcript of SEQ ID NO.2 comprising exon 20x (SEQ ID NO.10). In some embodiments, the ASOs disclosed herein target a SCN1A pre-mRNA sequence (SEQ ID NO. 2 or 12).
  • the ASO targets a SCN1A pre-mRNA sequence comprising an NIE (SEQ ID NO.10 or 20). In some embodiments, the ASO targets a SCN1A pre-mRNA sequence according to any one of SEQ ID NOs: 7-10 or 17-20. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 21-67. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 68-114. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 115- 209. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 210-256.
  • the ASO has a sequence according to any one of SEQ ID NOs: 257-303. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 304-341. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 342-379. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 380-1099. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 304-1099. In some embodiments, the ASO has a sequence according to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the SCN1A NIE containing pre-mRNA transcript is encoded by a genetic sequence with at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to SEQ ID NO.: 1 or 11.
  • the SCN1A NIE pre-mRNA transcript comprises a WSGR Docket No.47991-744.601 sequence with at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of SEQ ID NOs.: 2-10 and 12-20.
  • the ASO targets exon 20 of a SCN1A NIE containing pre-mRNA comprising NIE exon 20x. In some embodiments, the ASO targets an exon 21 sequence downstream (or 3’) of NIE exon 20x. In some embodiments, the ASO targets a sequence about 4 to about 300 nucleotides upstream (or 5’) from the 5’ end of exon 20x. In some embodiments, the ASO targets a sequence about 4 to about 300 nucleotides downstream (or 3’) from the 3’ end of exon 20x. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 21-67.
  • the ASO has a sequence according to any one of SEQ ID NOs: 210-256. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 380-1099. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 304-1099. In some embodiments, the ASO has a sequence according to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B. [0635] In some embodiments, the ASO targets a sequence upstream from the 5’ end of an NIE.
  • ASOs targeting a sequence upstream from the 5’ end of an NIE can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 21-38.
  • ASOs targeting a sequence upstream from the 5’ end of an NIE can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 68-85.
  • the ASOs target a sequence containing a exon- intron boundary (or junction).
  • ASOs targeting a sequence containing an exon-intron boundary can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 39-41, 51, 52, 228-230, 240, or 241.
  • ASOs targeting a sequence containing an exon-intron boundary can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 86-88 and 98-99.
  • the ASOs target a sequence downstream from the 3’ end of an NIE.
  • ASOs targeting a sequence down-stream from the 3’ end of an NIE can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 53-67.
  • ASOs targeting a sequence downstream from the 3’ end of an NIE can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 100-114.
  • ASOs target a sequence within an NIE.
  • ASOs targeting a sequence within an NIE can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 42-50, or 231-239.
  • ASOs targeting a sequence within an NIE e.g., exon 20x in human SCN1A, or exon 21x in mouse Scn1a
  • the ASO targets exon 20x in a SCN1A NIE containing pre-mRNA comprising exon 20x.
  • the ASO targets an exon 20x sequence downstream (or 3’) WSGR Docket No.47991-744.601 from the 5’ end of the exon 20x of a SCN1A pre-mRNA.
  • the ASO targets an exon 20x sequence upstream (or 5’) from the 3’ end of the exon 20x of a SCN1A pre-mRNA.
  • the SCN1A NIE containing pre-mRNA transcript comprises a sequence with at least about 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 2, 7-10, 12, and 17-20.
  • SCN1A NIE containing pre-mRNA transcript is encoded by a sequence with at least about 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to SEQ ID NOs: 1, 3-6, 11, and 13-16.
  • the targeted portion of the pre-mRNA containing an NMD exon and encoding NaV1.1 comprises a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to a region comprising at least 8 contiguous nucleic acids of SEQ ID NOs: 2, 7-10, 12, and 17-20.
  • the ASO targets a NIE containing pre-mRNA transcript.
  • the ASO targets a NIE containing pre-mRNA transcript comprising an NIE.
  • the ASO targets a NIE containing pre-mRNA transcript of comprising exon 20x.
  • the ASOs disclosed herein target a SCN1A pre-mRNA sequence.
  • the ASO targets a SCN1A pre-mRNA sequence comprising an NIE. In some embodiments, the ASO targets a SCN1A pre-mRNA sequence. In some embodiments, the ASO has a sequence according to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099. For another example, the ASOs comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the ASOs comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the ASO targets exon 20 of a SCN1A NIE containing pre-mRNA comprising NIE exon 20x.
  • the ASO targets an exon 21 sequence downstream (or 3') of NIE exon 20x.
  • the ASO targets a sequence about 4 to about 300 nucleotides upstream (or 5') from the 5' end of exon 20x.
  • the ASO targets a sequence about 4 to about 300 nucleotides downstream (or 3') from the 3' end of exon 20x. [0640] In some embodiments, the ASO targets a sequence upstream from the 5' end of an NIE. In some embodiments, the ASOs target a sequence containing an exon-intron boundary (or junction). In some embodiments, the ASOs target a sequence downstream from the 3' end of an NIE (e.g., exon 20x in human SCN1A, or exon 21x in mouse Scn1a). In some embodiments, ASOs target a sequence within an NIE.
  • an NIE e.g., exon 20x in human SCN1A, or exon 21x in mouse Scn1a.
  • the ASO targets exon 20x in a SCN1A NIE containing pre-mRNA comprising exon 20x. In some embodiments, the ASO targets an exon 20x sequence downstream (or 3') from the 5' end of the exon 20x of a SCN1A pre-mRNA. In some embodiments, the ASO targets an exon 20x sequence upstream (or 5') from the 3' end of the exon 20x of a SCN1A pre-mRNA.
  • the targeted portion of the SCN1A NIE containing pre-mRNA is in intron 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 (intron numbering corresponding to the mRNA sequence at NM_006920).
  • hybridization of an ASO WSGR Docket No.47991-744.601 to the targeted portion of the NIE pre-mRNA results in exon skipping of at least one of NIE within intron 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25, and subsequently increases NaV1.1 protein production.
  • hybridization of an ASO to the targeted portion of the NIE pre-mRNA inhibits or blocks exon skipping of at least one of NIE within intron 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25, and subsequently decreases NaV1.1 protein production.
  • the targeted portion of the SCN1A NIE containing pre-mRNA is in intron 20.
  • One of skill in the art can determine the corresponding intron number in any isoform based on an intron sequence provided herein or using the number provided in reference to the mRNA sequence at NM_006920, NM_001202435, NM_001165964, or NM_001165963.
  • compositions and methods comprising a therapeutic agent are provided to modulate protein expression level of SCN1A.
  • compositions and methods to modulate alternative splicing of SCNA1 pre-mRNA are provided herein.
  • compositions and methods to induce exon skipping in the splicing of SCN1A pre-mRNA e.g., to induce skipping of a pseudo-exon during splicing of SCN1A pre- mRNA.
  • therapeutic agents may be used to induce the inclusion of an exon in order to decrease the protein expression level.
  • a therapeutic agent disclosed herein is a small molecule, a polypeptide, or a polynucleic acid polymer. In some instances, the therapeutic agent is a small molecule. In some instances, the therapeutic agent is a polypeptide. In some instances, the therapeutic agent is a polynucleic acid polymer.
  • the therapeutic agent is a repressor agent. In additional cases, the therapeutic agent is an enhancer agent.
  • a therapeutic agent disclosed herein can be a NIE repressor agent.
  • a therapeutic agent may comprise a polynucleic acid polymer.
  • a method of treatment or prevention of a condition associated with a functional-NaV1.1 protein deficiency comprising administering a NIE repressor agent to a subject to increase levels of functional NaV1.1 protein, wherein the agent binds to a region of the pre-mRNA transcript to decrease inclusion of the NIE in the mature transcript.
  • a method of treatment or prevention of a condition associated with a functional-NaV1.1 protein deficiency comprising administering a NIE repressor agent to a subject to increase levels of functional NaV1.1 protein, wherein the agent binds to a region of an intron containing an NIE (e.g., intron 20 in human SCN1A gene) of the pre-mRNA transcript or to a NIE- activating regulatory sequence in the same intron.
  • an NIE e.g., intron 20 in human SCN1A gene
  • the sequence of the polynucleic acid polymer may be at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5% complementary to a target sequence of an mRNA transcript, e.g., a partially processed mRNA transcript.
  • the sequence of the polynucleic acid polymer may be 100% complementary to a target sequence of a pre-mRNA transcript.
  • the sequence of the polynucleic acid polymer may have 4 or fewer mismatches to a target sequence of the pre-mRNA transcript.
  • the sequence of the polynucleic acid polymer may have 3 or fewer mismatches to a target sequence of the pre-mRNA transcript.
  • the sequence of the polynucleic acid polymer may have 2 or fewer mismatches to a target sequence of the pre-mRNA transcript.
  • the sequence of the polynucleic acid polymer may have 1 or fewer mismatches to a tar-get sequence of the pre-mRNA transcript.
  • the sequence of the polynucleic acid polymer may have no mismatches to a target sequence of the pre-mRNA transcript.
  • the polynucleic acid polymer may specifically hybridize to a target sequence of the pre-mRNA transcript.
  • the polynucleic acid polymer may have 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5% or 100% sequence complementarity to a target sequence of the pre-mRNA transcript.
  • the hybridization may be under high stringent hybridization conditions.
  • the polynucleic acid polymer may have a sequence with at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5% sequence identity to a sequence selected from the group consisting of SEQ ID NOs: 21-67.
  • the polynucleic acid polymer may have a sequence with 100% sequence identity to a sequence selected from the group consisting of SEQ ID NOs: 21-67. In some instances, the polynucleic acid polymer may have a sequence with at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5% sequence identity to a sequence selected from the group consisting of SEQ ID NOs: 68-114. In some cases, the polynucleic acid polymer may have a sequence with 100% sequence identity to a sequence selected from the group consisting of SEQ ID NOs: 68-114.
  • the polynucleic acid polymer may have a sequence with at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5% sequence identity to a sequence selected from the group consisting of SEQ ID NOs: 21-67, 210-256 or 304-1099. In some cases, the polynucleic acid polymer may have a sequence with 100% sequence identity to a sequence selected from the group consisting of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the polynucleic acid polymer may have a sequence with 100% sequence identity to a sequence selected from the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the NIE repressor agent comprises a polynucleic acid polymer
  • the polynucleic acid polymer may be about 50 nucleotides in length.
  • the polynucleic acid polymer may be about 45 nucleotides in length.
  • the polynucleic acid polymer may be about 40 nucleotides in length.
  • the polynucleic acid polymer may be about 35 nucleotides in length.
  • the polynucleic acid polymer may be about 30 nucleotides in length.
  • the polynucleic acid polymer may be about 24 nucleotides in length.
  • the polynucleic acid polymer may be about 25 nucleotides in length.
  • the polynucleic acid polymer may be about 20 nucleotides in length.
  • the polynucleic acid polymer may be about 19 nucleotides in length.
  • the WSGR Docket No.47991-744.601 polynucleic acid polymer may be about 18 nucleotides in length.
  • the polynucleic acid polymer may be about 17 nucleotides in length.
  • the polynucleic acid polymer may be about 16 nucleotides in length.
  • the polynucleic acid polymer may be about 15 nucleotides in length.
  • the polynucleic acid polymer may be about 14 nucleotides in length.
  • the polynucleic acid polymer may be about 13 nucleotides in length.
  • the polynucleic acid polymer may be about 12 nucleotides in length.
  • the polynucleic acid polymer may be about 11 nucleotides in length.
  • the polynucleic acid polymer may be about 10 nucleotides in length.
  • the polynucleic acid polymer may be between about 10 and about 50 nucleotides in length.
  • the polynucleic acid polymer may be between about 10 and about 45 nucleotides in length.
  • the polynucleic acid polymer may be between about 10 and about 40 nucleotides in length.
  • the polynucleic acid polymer may be between about 10 and about 35 nucleotides in length.
  • the polynucleic acid polymer may be between about 10 and about 30 nucleotides in length.
  • the polynucleic acid polymer may be between about 10 and about 25 nucleotides in length.
  • the polynucleic acid polymer may be between about 10 and about 20 nucleotides in length.
  • the polynucleic acid polymer may be between about 15 and about 25 nucleotides in length.
  • the polynucleic acid polymer may be between about 15 and about 30 nucleotides in length.
  • the polynucleic acid polymer may be between about 12 and about 30 nucleotides in length.
  • the sequence of the polynucleic acid polymer may be at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5% complementary to a target sequence of an mRNA transcript, e.g., a partially processed mRNA transcript.
  • the sequence of the polynucleic acid polymer may be 100% complementary to a target sequence of a pre-mRNA transcript.
  • exon 20x in human SCN1A gene is equivalent to exon 21x in mouse Scn1a gene.
  • a method to identify or validate an NMD- inducing exon in the presence of an NMD inhibitor for example, cycloheximide.
  • an NMD inhibitor for example, cycloheximide.
  • references to sequence identity may be determined by BLAST sequence alignment using standard/default parameters.
  • the sequence may have 99% identity and still function according to the present disclosure.
  • the sequence may have 98% identity and still function according to the present disclosure.
  • the sequence may have 95% identity and still function according to the present disclosure.
  • the sequence may have 90% identity and still function according to the present disclosure.
  • Mode of Action [0657]
  • the reduced expression or function of NaV1.1 protein is associated with an altered splicing of a nonsense-mediated RNA decay-inducing exon (NMD exon) from a pre-mRNA that contains the NMD exon and that encodes NaV1.1 protein.
  • NMD exon nonsense-mediated RNA decay-inducing exon
  • the reduced expression or function of NaV1.1 protein can be associated with a reduced splicing of a nonsense-mediated RNA decay- WSGR Docket No.47991-744.601 inducing exon (NMD exon) from a pre-mRNA that contains the NMD exon and that encodes NaV1.1 protein.
  • NMD exon inducing exon
  • the ASO as described herein promotes exclusion of the NMD exon from a pre-mRNA that contains the NMD exon and that encodes NaV1.1 protein.
  • the ASO as described herein promotes exclusion of the NMD exon from a pre-mRNA that contains the NMD exon and that encodes NaV1.1 protein by at least 5%, 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%, at least 100%, at least 200%, at least 300%, at least 400%, at least 500%, at least 600%, at least 700%, at least 800%, at least 900%, at least 1000%, at least 2000%, at least 3000%, at least 4000%, at least 5000%, at least 6000%, at least 7000%, at least 8000%, at least 9000%, at least 10000%, at least 20000%, at least 30000%, at least 40000%, at least 50000%, at least 60000%, at least 70000%, at least 80000%, at least 90000%, or at least 100000% as compared to an untreated control cell, tissue or subject, or
  • the ASO as described herein promotes exclusion of the NMD exon a pre-mRNA that contains the NMD exon and that encodes Na V 1.1 protein by at least 2 fold, at least 3 fold, at least 4 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 25 fold, at least 30 fold, at least 40 fold, at least 50 fold, at least 60 fold, at least 70 fold, at least 80 fold, at least 90 fold, at least 100 fold, at least 200 fold, at least 300 fold, at least 400 fold, at least 500 fold, at least 600 fold, at least 700 fold, at least 800 fold, at least 900 fold, at least 1000 fold, at least 2000 fold, at least 3000 fold, at least 4000 fold, at least 5000 fold, at least 6000 fold, at least 7000 fold, at least 8000 fold, at least 9000 fold, or at least 10000 fold as compared to an untreated control cell, tissue or subject, or compared to the corresponding activity in the same type of cell, tissue or subject before treatment with the
  • the ASO as described herein binds to a targeted portion of a pre-mRNA that contains a nonsense-mediated RNA decay-inducing exon and that encodes Na V 1.1 protein.
  • the ASO as described herein promotes exclusion of a nonsense-mediated mRNA decay-inducing exon (NMD exon) from a pre-mRNA that contains the NMD exon and that encodes Na V 1.1 protein.
  • NMD exon a nonsense-mediated mRNA decay-inducing exon
  • the ASO as described herein promotes exclusion of an NMD exon from a pre-mRNA that contains the NMD exon and that encodes NaV1.1 protein by at least 5%, 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%, at least 100%, at least 200%, at least 300%, at least 400%, at least 500%, at least 600%, at least 700%, at least 800%, at least 900%, at least 1000%, at least 2000%, at least 3000%, at least 4000%, at least 5000%, at least 6000%, at least 7000%, at least 8000%, at least 9000%, at least 10000%, at least 20000%, at least 30000%, at least 40000%, at least 50000%, at least 60000%, at least 70000%, at least 80000%, at least 90000%, or at least 100000% as compared to an untreated control cell, tissue or subject, or
  • the ASO as WSGR Docket No.47991-744.601 described herein promotes exclusion of an NMD exon from a pre-mRNA that contains the NMD exon and that encodes NaV1.1 protein by at least 2 fold, at least 3 fold, at least 4 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 25 fold, at least 30 fold, at least 40 fold, at least 50 fold, at least 60 fold, at least 70 fold, at least 80 fold, at least 90 fold, at least 100 fold, at least 200 fold, at least 300 fold, at least 400 fold, at least 500 fold, at least 600 fold, at least 700 fold, at least 800 fold, at least 900 fold, at least 1000 fold, at least 2000 fold, at least 3000 fold, at least 4000 fold, at least 5000 fold, at least 6000 fold, at least 7000 fold, at least 8000 fold, at least 9000 fold, or at least 10000 fold as compared to an untreated control cell, tissue or subject, or compared to the corresponding
  • the ASO as described herein increases a level of processed mRNA encoding the NaV1.1 protein in a cell having a pre-mRNA that contains an NMD exon and that encodes Na V 1.1 protein when the ASO as described herein is introduced into the cell.
  • the ASO as described herein when the ASO as described herein is introduced into the cell, the ASO as described herein increases a level of processed mRNA encoding the Na V 1.1 protein in a cell having a pre- mRNA that contains an NMD exon and that encodes Na V 1.1 protein by at least 5%, 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%, at least 100%, at least 200%, at least 300%, at least 400%, at least 500%, at least 600%, at least 700%, at least 800%, at least 900%, at least 1000%, at least 2000%, at least 3000%, at least 4000%, at least 5000%, at least 6000%, at least 7000%, at least 8000%, at least 9000%, at least 10000%, at least 20000%, at least 30000%, at least 40000%, at least 50000%, at least 60000%, at least 70000%, at least 80000%,
  • the ASO as described herein when the ASO as described herein is introduced into the cell, the ASO as described herein increases a level of processed mRNA encoding the Na V 1.1 protein in a cell having a pre-mRNA that contains an NMD exon and that encodes Na V 1.1 protein by at least 2 fold, at least 3 fold, at least 4 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 25 fold, at least 30 fold, at least 40 fold, at least 50 fold, at least 60 fold, at least 70 fold, at least 80 fold, at least 90 fold, at least 100 fold, at least 200 fold, at least 300 fold, at least 400 fold, at least 500 fold, at least 600 fold, at least 700 fold, at least 800 fold, at least 900 fold, at least 1000 fold, at least 2000 fold, at least 3000 fold, at least 4000 fold, at least 5000 fold, at least 6000 fold, at least 7000 fold, at least 8000 fold, at least 9000 fold, or at least 10000 fold as compared to an un
  • the ASO as described herein increases a level of the NaV1.1 protein in a cell having a pre-mRNA that contains an NMD exon and that encodes NaV1.1 protein when the ASO as described herein is introduced into the cell.
  • the ASO as described herein when the ASO as described herein is introduced into the cell, the ASO as described herein increases a level of the NaV1.1 protein in a cell having a pre-mRNA that contains an WSGR Docket No.47991-744.601 NMD exon and that encodes NaV1.1 protein by at least 5%, 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%, at least 100%, at least 200%, at least 300%, at least 400%, at least 500%, at least 600%, at least 700%, at least 800%, at least 900%, at least 1000%, at least 2000%, at least 3000%, at least 4000%, at least 5000%, at least 6000%, at least 7000%, at least 8000%, at least 9000%, at least 10000%, at least 20000%, at least 30000%, at least 40000%, at least 50000%, at least 60000%, at least 70000%, at least
  • the ASO as described herein when the ASO as described herein is introduced into the cell, the ASO as described herein increases a level of the NaV1.1 protein in a cell having a pre-mRNA that contains an NMD exon and that encodes NaV1.1 protein by at least 2 fold, at least 3 fold, at least 4 fold, at least 5 fold, at least 10 fold, at least 20 fold, at least 25 fold, at least 30 fold, at least 40 fold, at least 50 fold, at least 60 fold, at least 70 fold, at least 80 fold, at least 90 fold, at least 100 fold, at least 200 fold, at least 300 fold, at least 400 fold, at least 500 fold, at least 600 fold, at least 700 fold, at least 800 fold, at least 900 fold, at least 1000 fold, at least 2000 fold, at least 3000 fold, at least 4000 fold, at least 5000 fold, at least 6000 fold, at least 7000 fold, at least 8000 fold, at least 9000 fold, or at least 10000 fold as compared to an untreated control cell, tissue or subject, or
  • the targeted portion is within an intron sequence flanking the NMD exon.
  • the targeted portion comprises at least one nucleotide of the NMD exon.
  • the targeted portion comprises at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, or 49 nucleotide of the NMD exon.
  • the targeted portion comprises at most 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, or 49 nucleotide of the NMD exon.
  • the targeted portion is within the NMD exon.
  • the NMD exon comprises a sequence with at least 80%, at least 90%, or 100% sequence identity to a sequence selected from the group consisting of the sequences listed in Table 1 or Table 2.
  • the NMD exon comprises a sequence selected from the group consisting of the sequences listed in Table 1 or Table 2.
  • the pre-mRNA comprises a sequence with at least about 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to a sequence selected from the group consisting of the sequences listed in Table 1 or Table 2.
  • the pre-mRNA is encoded by a genetic sequence with at least about 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to a sequence selected from the group consisting of the sequences listed in Table 1 or Table 2.
  • spliceosome a large and highly dynamic RNA-protein complex termed the spliceosome, which orchestrates complex interactions between primary transcripts, small nuclear RNAs (snRNAs) and a large number of proteins.
  • Spliceosomes assemble ad hoc on each intron in an ordered manner, starting with recognition of the 5' splice site (5'ss) by U1 snRNA or the 3' splice site (3'ss) by the U2 pathway, which involves binding of the U2 auxiliary factor (U2AF) to the 3'ss region to facilitate U2 binding to the branch point sequence (BPS).
  • 5'ss 5' splice site
  • U1 snRNA the 3' splice site (3'ss) by the U2 pathway
  • U2AF U2 auxiliary factor
  • U2AF is a stable heterodimer composed of a U2AF2-encoded 65-kD subunit (U2AF65), which binds the polypyrimidine tract (PPT), and a U2AF1-encoded 35-kD subunit (U2AF35), which interacts with highly conserved AG dinucleotides at 3'ss and stabilizes U2AF65 binding.
  • U2AF65 U2AF2-encoded 65-kD subunit
  • PPT polypyrimidine tract
  • U2AF35 U2AF1-encoded 35-kD subunit
  • accurate splicing requires auxiliary sequences or structures that activate or repress splice site recognition, known as intronic or exonic splicing enhancers or silencers.
  • ESRs auxiliary exonic and intronic splicing regulatory elements
  • these functional elements are classified as either exonic or intronic splicing enhancers (ESEs or ISEs) or silencers (ESSs or ISSs) based on their ability to stimulate or inhibit splicing, respectively.
  • EESs or ISEs exonic or intronic splicing enhancers
  • ESSs or ISSs silencers
  • auxiliary cis-acting elements may act by influencing the kinetics of spliceosome assembly, such as the arrangement of the complex between U1 snRNP and the 5'ss, it seems very likely that many elements function in concert with trans-acting RNA-binding proteins (RBPs).
  • SR proteins serine- and arginine-rich family of RBPs
  • SR proteins promote exon recognition by recruiting components of the pre-spliceosome to adjacent splice sites or by antagonizing the effects of ESSs in the vicinity.
  • the repressive effects of ESSs can be mediated by members of the heterogeneous nuclear ribonucleoprotein (hnRNP) family and can alter recruitment of core splicing factors to adjacent splice sites.
  • hnRNP heterogeneous nuclear ribonucleoprotein
  • silencer elements are suggested to have a role in repression of pseudo-exons, sets of decoy intronic splice sites with the typical spacing of an exon but without a functional open reading frame.
  • ESEs and ESSs in cooperation with their cognate trans-acting RBPs, represent important components in a set of splicing controls that specify how, where and when mRNAs are assembled from their precursors.
  • the sequences marking the exon-intron boundaries are degenerate signals of varying strengths that can occur at high frequency within human genes. In multi-exon genes, different pairs of splice sites can be linked together in many different combinations, creating a diverse array of transcripts from a single gene.
  • alternative pre-mRNA splicing This is commonly referred to as alternative pre-mRNA splicing.
  • most mRNA WSGR Docket No.47991-744.601 isoforms produced by alternative splicing can be exported from the nucleus and translated into functional polypeptides, different mRNA isoforms from a single gene can vary greatly in their translation efficiency.
  • Those mRNA isoforms with premature termination codons (PTCs) at least 50 bp upstream of an exon junction complex are likely to be targeted for degradation by the nonsense-mediated mRNA decay (NMD) pathway.
  • PTCs premature termination codons
  • Mutations in traditional (BPS/PPT/3'ss/5'ss) and auxiliary splicing motifs can cause aberrant splicing, such as exon skipping or cryptic (or pseudo-) exon inclusion or splice-site activation, and contribute significantly to human morbidity and mortality.
  • Both aberrant and alternative splicing patterns can be influenced by natural DNA variants in exons and introns.
  • exon-intron boundaries can occur at any of the three positions of a codon, it is clear that only a subset of alternative splicing events can maintain the canonical open reading frame. For example, only exons that are evenly divisible by 3 can be skipped or included in the mRNA without any alteration of reading frame.
  • NMD is a translation-coupled mechanism that eliminates mRNAs containing PTCs. NMD can function as a surveillance pathway that exists in all eukaryotes. NMD can reduce errors in gene expression by eliminating mRNA transcripts that contain premature stop codons. Translation of these aberrant mRNAs could, in some cases, lead to deleterious gain-of-function or dominant-negative activity of the resulting proteins.
  • NMD NMD targets not only transcripts with PTCs but also a broad array of mRNA isoforms expressed from many endogenous genes, suggesting that NMD is a master regulator that drives both fine and coarse adjustments in steady-state RNA levels in the cell.
  • NMD-inducing exon NIE is an exon or a pseudo-exon that is a region within an intron and can activate the NMD pathway if included in a mature RNA transcript.
  • the intron containing an NIE is usually spliced out, but the intron or a portion thereof (e.g., NIE) can be retained during alternative or aberrant splicing events.
  • Mature mRNA transcripts containing such an NIE can be non-productive due to frame shift which induce NMD pathway. Inclusion of a NIE in mature RNA transcripts can downregulate gene expression.
  • mRNA transcripts, such as a pre-mRNA transcript, containing an NIE can be referred as “NIE containing mRNA” or “NMD exon mRNA” in the current disclosure.
  • NIE containing mRNA or “NMD exon mRNA” in the current disclosure.
  • Cryptic (or pseudo-splice sites) have the same splicing recognition sequences as genuine splice sites but are not used in the splicing reactions. They outnumber genuine splice sites in the human genome by an order of a magnitude and are normally repressed by thus far poorly understood molecular mechanisms.
  • Cryptic 5' splice sites have the consensus NNN/GUNNNN or NNN/GCNNNN where N is any nucleotide and/is the exon-intron boundary.
  • Cryptic 3' splice sites have the consensus NAG/N. Their activation is positively influenced by surrounding nucleotides that make them more similar to the optimal consensus of authentic splice sites, namely MAG/GURAGU and YAG/G, respectively, where M is C or A, R is G or A, and Y is C or U.
  • Splice sites and their regulatory sequences can be readily identified by a skilled person using suitable algorithms publicly available, listed for example in Kralovicova, J. and Vorechovsky, I.
  • cryptic splice sites or splicing regulatory sequences may compete for RNA-binding proteins such as U2AF with a splice site of the NIE.
  • an agent may bind to the cryptic splice site or splicing regulatory sequences to prevent the binding of RNA-binding proteins and thereby favoring utilization of the NIE splice sites.
  • the cryptic splice site may not comprise the 5' or 3' splice site of the NIE.
  • the cryptic splice site may be at least 10 nucleotides upstream of the NIE 5' splice site.
  • the cryptic splice site may be at least 20 nucleotides upstream of the NIE 5' splice site.
  • the cryptic splice site may be at least 50 nucleotides upstream of the NIE 5' splice site.
  • the cryptic splice site may be at least 100 nucleotides upstream of the NIE 5' splice site.
  • the cryptic splice site may be at least 200 nucleotides upstream of the NIE 5' splice site.
  • the cryptic splice site may be at least 10 nucleotides downstream of the NIE 3' splice site.
  • the cryptic splice site may be at least 20 nucleotides downstream of the NIE 3' splice site.
  • the cryptic splice site may be at least 50 nucleotides downstream of the NIE 3' splice site.
  • the cryptic splice site may be at least 100 nucleotides downstream of the NIE 3' splice site.
  • the cryptic splice site may be at least 200 nucleotides downstream of the NIE 3' splice site.
  • the methods of the present disclosure exploit the presence of NIE in the pre-mRNA transcribed from the SCN1A gene. Splicing of the identified SCN1A NIE pre-mRNA species to produce functional mature SCN1A mRNA can be induced using a therapeutic agent such as an ASO that stimulates exon skipping of an NIE. Induction of exon skipping can result in inhibition of an NMD pathway.
  • the resulting mature SCN1A mRNA can be translated normally without activating NMD pathway, thereby increasing the amount of Na V 1.1 (also termed “Na V 1.1 protein” herein) in the patient’s cells and alleviating symptoms of a condition associated with SCN1A deficiency, such as Dravet Syndrome (DS); Epilepsy, generalized, with febrile seizures plus, type 2; Febrile seizures, familial, 3A; Autism; Epileptic encephalopathy, early infantile, 13; Sick sinus syndrome 1; Alzheimer’s disease; or SUDEP.
  • DS Dravet Syndrome
  • Epilepsy generalized, with febrile seizures plus, type 2; Febrile seizures, familial, 3A; Autism
  • Epileptic encephalopathy early infantile, 13
  • Sick sinus syndrome 1 Alzheimer’s disease
  • Alzheimer’s disease or SUDEP.
  • the reduction/correction may be relative to the level of NIE inclusion in the subject without treatment, or relative to the amount of NIE inclusion in a population of similar subjects.
  • the reduction/correction may be at least 10% less NIE inclusion relative to the average subject, or the subject prior to treatment.
  • the reduction may be at least 20% less NIE inclusion relative to an average subject, or the subject prior to treatment.
  • the reduction may be at least 40% less NIE inclusion relative to an average subject, or the subject prior to treatment.
  • the reduction may be at least 50% less NIE inclusion relative to an average subject, or the subject prior to treatment.
  • the reduction may be at least 60% less NIE inclusion relative to an average subject, or the subject prior to treatment.
  • the reduction may be at least 80% less NIE WSGR Docket No.47991-744.601 inclusion relative to an average subject, or the subject prior to treatment.
  • the reduction may be at least 90% less NIE inclusion relative to an average subject, or the subject prior to treatment.
  • the increase may be clinically significant.
  • the increase may be relative to the level of active-NaV1.1 protein in the subject without treatment, or relative to the amount of active-NaV1.1 protein in a population of similar subjects.
  • the increase may be at least 10% more active-NaV1.1 protein relative to the average subject, or the subject prior to treatment.
  • the increase may be at least 20% more active-NaV1.1 protein relative to the average subject, or the subject prior to treatment.
  • the increase may be at least 40% more active-NaV1.1 protein relative to the average subject, or the subject prior to treatment.
  • the increase may be at least 50% more active-NaV1.1 protein relative to the average subject, or the subject prior to treatment.
  • the increase may be at least 80% more active-NaV1.1 protein relative to the average subject, or the subject prior to treatment.
  • the increase may be at least 100% more active-Na V 1.1 protein relative to the average subject, or the subject prior to treatment.
  • the increase may be at least 200% more active-Na V 1.1 protein relative to the average subject, or the subject prior to treatment.
  • the increase may be at least 500% more active- Na V 1.1 protein relative to the average subject, or the subject prior to treatment.
  • a “NIE containing pre-mRNA” is a pre-mRNA transcript that contains at least one pseudo-exon. Alternative or aberrant splicing can result in inclusion of the at least one pseudo-exon in the mature mRNA transcripts.
  • the terms “mature mRNA,” and “fully spliced mRNA,” are used interchangeably here-in to describe a fully processed mRNA. Inclusion of the at least one pseudo-exon can be non-productive mRNA and lead to NMD of the mature mRNA. NIE containing mature mRNA may sometimes lead to aberrant protein expression.
  • One of skill in the art also can determine the sequences of flanking exons in any SCN1A isoform for targeting using the methods of the disclosure, based on an intron sequence provided herein or using the intron number provided in reference to the mRNA sequence at NM_006920, NM_001202435, NM_001165964, or NM_001165963.
  • the methods and compositions of the present disclosure are used to modulate, e.g., increase or decrease, the expression of SCN1A by inducing or inhibiting exon skipping of a pseudo-exon of an SCN1A NIE containing pre-mRNA.
  • the pseudo-exon is a sequence within any of introns 1-25. In some embodiments, the pseudo-exon is a sequence within any of introns 2, 4, 6, 13, 14, 15, 16, 17, 18, 20, 21, 22, 23, 24, and 25. In some embodiments, the pseudo-exon is a sequence within any of introns 15, 18, and 19. In some embodiments, the pseudo-exon can be any SCN1A intron or a portion thereof. In some embodiments, the pseudo-exon is within intron 20.
  • the SCN1A intron numbering used herein corresponds to the mRNA sequence at NM_006920. It is understood that the intron numbering may change in reference to a different SCN1A isoform sequence.
  • the included pseudo-exon is the most abundant pseudo-exon in a population of NIE containing pre-mRNAs transcribed from the gene encoding the target protein in a cell. In some embodiments, the included pseudo-exon is the most abundant pseudo-exon in a population of WSGR Docket No.47991-744.601 NIE containing pre-mRNAs transcribed from the gene encoding the target protein in a cell, where-in the population of NIE containing pre-mRNAs comprises two or more included pseudo-exons.
  • an antisense oligomer targeted to the most abundant pseudo-exon in the population of NIE containing pre-mRNAs encoding the target protein induces exon skipping of one or two or more pseudo- exons in the population, including the pseudo-exon to which the antisense oligomer is targeted or binds.
  • the targeted region is in a pseudo-exon that is the most abundant pseudo-exon in a NIE containing pre-mRNA encoding the NaV1.1 protein.
  • the degree of exon inclusion can be expressed as percent exon inclusion, e.g., the percentage of transcripts in which a given pseudo-exon is included.
  • an included pseudo-exon is an exon that is identified as an included pseudo-exon based on a determination of at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, or at least about 50%, inclusion.
  • a included pseudo-exon is an exon that is identified as a included pseudo-exon based on a determination of about 5% to about 100%, about 5% to about 95%, about 5% to about 90%, about 5% to about 85%, about 5% to about 80%, about 5% to about 75%, about 5% to about 70%, about 5% to about 65%, about 5% to about 60%, about 5% to about 55%, about 5% to about 50%, about 5% to about 45%, about 5% to about 40%, about 5% to about 35%, about 5% to about 30%, about 5% to about 25%, about 5% to about 20%, about 5% to about 15%, about 10% to about 100%, about 10% to about 95%, about 10% to about 90%, about 10% to about 85%, about 10% to about 80%, about 10% to about 75%, about 10% to about 70%, about 10% to about 65%, about 10% to about 60%, about 10% to about 55%, about 10% to about 50%, about 10% to about 45%, about 10% to about 40%, about 10% to about 100%,
  • ENCODE data (described by, e.g., Tilgner, et al., 2012, “Deep sequencing of subcellular RNA fractions shows splicing to be predominantly co- WSGR Docket No.47991-744.601 transcriptional in the human genome but inefficient for lncRNAs,” Genome Research 22(9):1616-25, of which entire content is incorporated herein by reference) can be used to aid in identifying exon inclusion.
  • contacting cells with an ASO that is complementary to a targeted portion of a SCN1A pre-mRNA transcript results in an increase in the amount of NaV1.1 protein produced by at least 10, 20, 30, 40, 50, 60, 80, 100, 150, 200, 250, 300, 350, 400, 450, 500, or 1000%, compared to the amount of the protein produced by a cell in the absence of the ASO/absence of treatment.
  • the total amount of NaV1.1 protein produced by the cell to which the antisense oligomer is contacted is increased about 1.1 to about 10-fold, about 1.5 to about 10-fold, about 2 to about 10-fold, about 3 to about 10-fold, about 4 to about 10-fold, about 1.1 to about 5-fold, about 1.1 to about 6-fold, about 1.1 to about 7-fold, about 1.1 to about 8-fold, about 1.1 to about 9-fold, about 2 to about 5-fold, about 2 to about 6-fold, about 2 to about 7-fold, about 2 to about 8-fold, about 2 to about 9-fold, about 3 to about 6-fold, about 3 to about 7-fold, about 3 to about 8-fold, about 3 to about 9-fold, about 4 to about 7-fold, about 4 to about 8-fold, about 4 to about 9-fold, at least about 1.1-fold, at least about 1.5-fold, at least about 2-fold, at least about 2.5-fold, at least about 3-fold, at least about 3.5-fold, at least about 4- fold, at least about 1.1 to about
  • a control compound can be, for example, an oligonucleotide that is not complementary to a targeted portion of the pre-mRNA.
  • contacting cells with an ASO that is complementary to a targeted portion of a SCN1A pre-mRNA transcript results in a decrease in the amount of Na V 1.1 protein produced by at least 10, 20, 30, 40, 50, 60, 80, 100, 150, 200, 250, 300, 350, 400, 450, 500, or 1000%, compared to the amount of the protein produced by a cell in the absence of the ASO/absence of treatment.
  • the total amount of Na V 1.1 protein produced by the cell to which the antisense oligomer is contacted is decreased about 1.1 to about 10-fold, about 1.5 to about 10-fold, about 2 to about 10-fold, about 3 to about 10-fold, about 4 to about 10-fold, about 1.1 to about 5-fold, about 1.1 to about 6-fold, about 1.1 to about 7-fold, about 1.1 to about 8-fold, about 1.1 to about 9-fold, about 2 to about 5-fold, about 2 to about 6-fold, about 2 to about 7-fold, about 2 to about 8-fold, about 2 to about 9-fold, about 3 to about 6-fold, about 3 to about 7-fold, about 3 to about 8-fold, about 3 to about 9-fold, about 4 to about 7-fold, about 4 to about 8-fold, about 4 to about 9-fold, at least about 1.1-fold, at least about 1.5-fold, at least about 2-fold, at least about 2.5-fold, at least about 3-fold, at least about 3.5-fold, at least about 4- fold, at least about 1.1 to
  • a control compound can be, for example, an oligonucleotide that is not complementary to a targeted portion of the pre-mRNA.
  • the amount of mRNA encoding NaV1.1 protein, or the mature mRNA encoding the NaV1.1 protein is increased by at least 10, 20, 30, 40, 50, 60, 80, 100, 150, 200, 250, 300, 350, 400, 450, 500, or 1000%, compared to the amount of the protein produced by a cell in the absence of the ASO/absence of treatment.
  • the WSGR Docket No.47991-744.601 total amount of the mRNA encoding NaV1.1 protein, or the mature mRNA encoding NaV1.1 protein produced in the cell to which the antisense oligomer is contacted is increased about 1.1 to about 10-fold, about 1.5 to about 10-fold, about 2 to about 10-fold, about 3 to about 10-fold, about 4 to about 10-fold, about 1.1 to about 5-fold, about 1.1 to about 6-fold, about 1.1 to about 7-fold, about 1.1 to about 8-fold, about 1.1 to about 9-fold, about 2 to about 5-fold, about 2 to about 6-fold, about 2 to about 7-fold, about 2 to about 8-fold, about 2 to about 9-fold, about 3 to about 6-fold, about 3 to about 7-fold, about 3 to about 8-fold, about 3 to about 9-fold, about 4 to about 7-fold, about 4 to about 8-fold, about 4 to about 9- fold, at least about 1.1-fold, at least about 1.5-fold,
  • a control compound can be, for example, an oligonucleotide that is not complementary to a targeted portion of the SCN1A NIE containing pre-mRNA.
  • contacting cells with an ASO that is complementary to a targeted portion of a SCN1A pre-mRNA transcript results in a decrease in the amount of mRNA encoding SCN1A, including the mature mRNA encoding the target protein.
  • the amount of mRNA encoding Na V 1.1 protein, or the mature mRNA encoding the Na V 1.1 protein is decreased by at least 10, 20, 30, 40, 50, 60, 80, 100, 150, 200, 250, 300, 350, 400, 450, 500, or 1000%, compared to the amount of the protein produced by a cell in the absence of the ASO/absence of treatment.
  • the total amount of the mRNA encoding Na V 1.1 protein, or the mature mRNA encoding Na V 1.1 protein produced in the cell to which the antisense oligomer is contacted is decreased about 1.1 to about 10-fold, about 1.5 to about 10-fold, about 2 to about 10-fold, about 3 to about 10-fold, about 4 to about 10-fold, about 1.1 to about 5-fold, about 1.1 to about 6-fold, about 1.1 to about 7-fold, about 1.1 to about 8-fold, about 1.1 to about 9-fold, about 2 to about 5-fold, about 2 to about 6-fold, about 2 to about 7-fold, about 2 to about 8-fold, about 2 to about 9-fold, about 3 to about 6-fold, about 3 to about 7-fold, about 3 to about 8-fold, about 3 to about 9-fold, about 4 to about 7-fold, about 4 to about 8-fold, about 4 to about 9- fold, at least about 1.1-fold, at least about 1.5-fold, at least about 2-fold, at least about 2.5-fold,
  • a control compound can be, for example, an oligonucleotide that is not complementary to a targeted portion of the SCN1A NIE containing pre-mRNA.
  • the NIE can be in any length. In some embodiments, the NIE comprises a full sequence of an intron, in which case, it can be referred to as intron retention. In some embodiments, the NIE can be a portion of the intron. In some embodiments, the NIE can be a 5' end portion of an intron including a 5'ss sequence. In some embodiments, the NIE can be a 3' end portion of an intron including a 3'ss sequence. In some embodiments, the NIE can be a portion within an intron without inclusion of a 5'ss sequence.
  • the NIE can be a portion within an intron without inclusion of a 3'ss sequence. In some embodiments, the NIE can be a portion within an intron without inclusion of either a 5'ss or a 3'ss sequence.
  • the NIE can be from 5 nucleotides to 10 nucleotides in length, from 10 WSGR Docket No.47991-744.601 nucleotides to 15 nucleotides in length, from 15 nucleotides to 20 nucleotides in length, from 20 nucleotides to 25 nucleotides in length, from 25 nucleotides to 30 nucleotides in length, from 30 nucleotides to 35 nucleotides in length, from 35 nucleotides to 40 nucleotides in length, from 40 nucleotides to 45 nucleotides in length, from 45 nucleotides to 50 nucleotides in length, from 50 nucleotides to 55 nucleotides in length, from 55 nucleotides to 60 nucleotides in length, from 60 nucleotides to 65 nucleotides in length, from 65 nucleotides to 70 nucleotides in length, from 70 nucleotides to 75
  • the NIE can be at least 10 nucleotides, at least 20 nucleotides, at least 30 nucleotides, at least 40 nucleotides, at least 50 nucleotides, at least 60 nucleoids, at least 70 nucleotides, at least 80 nucleotides in length, at least 90 nucleotides, or at least 100 nucleotides in length.
  • the NIE can be from 100 to 200 nucleotides in length, from 200 to 300 nucleotides in length, from 300 to 400 nucleotides in length, from 400 to 500 nucleotides in length, from 500 to 600 nucleotides in length, from 600 to 700 nucleotides in length, from 700 to 800 nucleotides in length, from 800 to 900 nucleotides in length, from 900 to 1,000 nucleotides in length. In some embodiments, the NIE may be longer than 1,000 nucleotides in length. [0696] Inclusion of a pseudo-exon can lead to a frameshift and the introduction of a premature termination codon (PIC) in the mature mRNA transcript rendering the transcript a target of NMD.
  • PIC premature termination codon
  • Mature mRNA transcript containing NIE can be non-productive mRNA transcript which does not lead to protein expression.
  • the PIC can be present in any position downstream of an NIE. In some embodiments, the PIC can be present in any exon downstream of an NIE. In some embodiments, the PIC can be present within the NIE. For example, inclusion of exon 20x in an mRNA transcript encoded by the SCN1A gene can induce a PIC in the mRNA transcript, e.g., a PIC in exon 21 of the mRNA transcript.
  • ASOs that when hybridized to a region of a pre-mRNA result in exon skipping (or enhanced splicing of the intron containing a NIE) and increased protein production may be tested in vivo using animal models, for example transgenic mouse models in which the full-length human gene has been knocked-in or in humanized mouse models of disease. Suitable routes for administration of ASOs may vary depending on the disease and/or the cell types to which delivery of the ASOs is desired. ASOs may be administered, for example, by intrathecal injection, intracerebroventricular injection, intraperitoneal injection, intramuscular injection, subcutaneous injection, intravitreal injection, or intravenous injection.
  • the cells, tissues, and/or organs of the model animals may be assessed to determine the effect of the ASO treatment by for example evaluating splicing (efficiency, rate, extent) and protein production by methods known in the art and described herein.
  • the animal models may also be any phenotypic or behavioral indication of the disease or disease severity.
  • the antisense oligonucleotide is administered with one or more agents capable of promoting penetration of the subject antisense oligonucleotide across the blood-brain barrier by any method known in the art. For example, delivery of agents by administration of an adenovirus vector to WSGR Docket No.47991-744.601 motor neurons in muscle tissue is described in U.S. Pat. No.
  • 6,632,427 “Adenoviral-vector-mediated gene transfer into medullary motor neurons,” incorporated herein by reference. Delivery of vectors directly to the brain, e.g., the striatum, the thalamus, the hippocampus, or the substantia nigra, is described, e.g., in U.S. Pat. No.6,756,523, “Adenovirus vectors for the transfer of foreign genes into cells of the central nervous system particularly in brain,” incorporated herein by reference. [0699] In embodiments, the antisense oligonucleotides are linked or conjugated with agents that provide desirable pharmaceutical or pharmacodynamic properties.
  • the antisense oligonucleotide is coupled to a substance, known in the art to promote penetration or transport across the blood-brain barrier, e.g., an antibody to the transferrin receptor.
  • the antisense oligonucleotide is linked with a viral vector, e.g., to render the antisense compound more effective or increase transport across the blood-brain barrier.
  • osmotic blood brain barrier disruption is assisted by infusion of sugars, e.g., meso erythritol, xylitol, D(+) galactose, D(+) lactose, D(+) xylose, dulcitol, myo- inositol, L(-) fructose, D(-) mannitol, D(+) glucose, D(+) arabinose, D(-) arabinose, cellobiose, D(+) maltose, D(+) raffinose, L(+) rhamnose, D(+) melibiose, D(-) ribose, adonitol, D(+) arabitol, L(-) arabitol, D(+) fucose, L(-) fucose, D(-) lyxose, L(+) lyxose, and L(-
  • an ASO of the disclosure is coupled to a dopamine reuptake inhibitor (DRI), a selective serotonin reuptake inhibitor (SSRI), a noradrenaline reuptake inhibitor (NRI), a norepinephrine- dopamine reuptake inhibitor (NDRI), and a serotonin-norepinephrine-dopamine reuptake inhibitor (SNDRI), using methods described in, e.g., U.S. Pat. No.9,193,969, incorporated herein by reference.
  • subjects treated using the methods and compositions are evaluated for improvement in condition using any methods known and described in the art.
  • the SCN1A NIE containing pre-mRNA transcript is encoded by a genetic sequence with at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of the sequences listed in Table 1 or Table 2.
  • the SCN1A NIE pre- mRNA transcript comprises a sequence with at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to any one of the sequences listed in Table 1 or Table 2.
  • the SCN1A NIE containing pre-mRNA transcript comprises a sequence with at least about 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of the sequences listed in Table 1 or Table 2.
  • SCN1A NIE containing pre-mRNA transcript is encoded by a sequence with at least about 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of the sequences listed in Table 1 or Table 2.
  • the targeted portion of the pre- WSGR Docket No.47991-744.601 mRNA containing an NMD exon and encoding NaV1.1 comprises a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to a region comprising at least 8 contiguous nucleic acids of any one of the sequences listed in Table 1 or Table 2.
  • the pre-mRNA transcript comprises a sequence with at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a pre-mRNA transcript of SCN1A pre-mRNA transcripts or a complement thereof described herein.
  • the targeted portion of the pre-mRNA selected from the group consisting of SCN1A pre-mRNAs comprises a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to a region comprising at least 8 contiguous nucleic acids of a sequence of the pre-mRNA transcripts of Table 1 or Table 2 or complements thereof.
  • the targeted portion of the pre-mRNA of SCN1A pre- mRNA comprises a sequence that is complementary to at least 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 contiguous nucleic acids of a sequence of Table 1 or Table 2 or a complement thereof.
  • the pre-mRNA transcript comprises a sequence with at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity to a pre-mRNA transcript of SCN1A pre-mRNA transcripts or a complement thereof described herein.
  • the targeted portion of the pre-mRNA selected from the group consisting of SCN1A pre-mRNAs comprises a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to a region comprising at least 8 contiguous nucleic acids of a sequence of the pre-mRNA transcripts of Table 1 or Table 2 or complements thereof.
  • the targeted portion of the pre-mRNA of SCN1A pre- mRNA comprises a sequence that is complementary to at least 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 contiguous nucleic acids of a sequence of Table 1 or Table 2 or a complement thereof.
  • the targeted portion of the SCN1A pre-mRNA comprises a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to a region comprising at least 8 contiguous nucleic acids of a sequence of sequence of Table 3 or complements thereof.
  • the targeted portion of the SCN1A pre-mRNA comprises a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to a region comprising at least 8 contiguous nucleic acids of a sequence selected from the group consisting of sequences listed in Table 2 or Table 3 or complements thereof.
  • the ASO has a sequence complementary to the targeted portion of the pre- mRNA containing an NMD exon according to any one of the sequences listed in Table 1 or Table 2.
  • the ASO targets a sequence upstream from the 5’ end of an NIE.
  • ASOs targeting a sequence upstream from the 5’ end of an NIE comprises a sequence that is at least about 80%, 85%, 90%, 95%, 97%, or 100% complementary to at least 8 contiguous nucleic acids of any one of the sequences listed in Table 1 or Table 2.
  • ASOs targeting a sequence upstream from the 5’ end of an NIE can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of the sequences listed in Table 1 or Table 2.
  • the ASOs target a sequence containing an exon-intron boundary (or junction).
  • ASOs targeting a sequence containing an exon-intron boundary can comprise a WSGR Docket No.47991-744.601 sequence that is at least about 80%, 85%, 90%, 95%, 97%, or 100% complementary to at least 8 contiguous nucleic acids of any one of the sequences listed in Table 1 or Table 2.
  • the ASOs target a sequence downstream from the 3’ end of an NIE.
  • ASOs targeting a sequence downstream from the 3’ end of an NIE can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of the sequences listed in Table 1 or Table 2.
  • ASOs targeting a sequence downstream from the 3’ end of an NIE can comprise a sequence with at least 80%, 85%, 90%, 95%, 97%, or 100% sequence identity to any one of the sequences listed in Table 1 or Table 2.
  • ASOs target a sequence within an NIE.
  • ASOs that specifically hybridize to different nucleotides within the target region of the pre- mRNA may be screened to identify or determine ASOs that improve the rate and/or extent of splicing of the target intron.
  • the ASO may block or interfere with the binding site(s) of a splicing repressor(s)/silencer. Any method known in the art may be used to identify (determine) an ASO that when hybridized to the target region of the exon results in the desired effect (e.g., pseudo-exon skipping, protein or functional RNA production).
  • a round of screening referred to as an ASO “walk” may be performed using ASOs that have been designed to hybridize to a target region of a pre-mRNA.
  • the ASOs used in the ASO walk can be tiled every 5 nucleotides from approximately 100 nucleotides upstream of the 3’ splice site of the included exon (e.g., a portion of sequence of the exon located upstream of the target/included exon) to approximately 100 nucleotides downstream of the 3’ splice site of the target/included exon and/or from approximately 100 nucleotides upstream of the 5’ splice site of the included exon to approximately 100 nucleotides downstream of the 5’ splice site of the target/included exon (e.g., a portion of sequence of the exon located downstream of the target/included exon).
  • a first ASO of 15 nucleotides in length may be designed to specifically hybridize to nucleotides +6 to +20 relative to the 3’ splice site of the target/included exon.
  • a second ASO may be designed to specifically hybridize to nucleotides +11 to +25 relative to the 3’ splice site of the target/included exon.
  • ASOs are designed as such spanning the target region of the pre-mRNA. In embodiments, the ASOs can be tiled more closely, e.g., every 1, 2, 3, or 4 nucleotides.
  • the ASOs can be tiled from 100 nucleotides downstream of the 5’ splice site, to 100 nucleotides upstream of the 3’ splice site. In some embodiments, the ASOs can be tiled from about 1,160 nucleotides upstream of the 3’ splice site, to about 500 nucleotides downstream of the 5’ splice site. In some embodiments, the ASOs can be tiled from about 500 nucleotides upstream of the 3’ splice site, to about 1,920 nucleotides downstream of the 3’ splice site.
  • One or more ASOs, or a control ASO are delivered, for example by transfection, into a disease- relevant cell line that expresses the target pre-mRNA (e.g., a NIE containing pre-mRNA described herein).
  • a disease- relevant cell line that expresses the target pre-mRNA (e.g., a NIE containing pre-mRNA described herein).
  • the exon skipping effects of each of the ASOs may be assessed by any method known in the art, for example by reverse transcriptase (RT)-PCR using primers that span the splice junction.
  • RT reverse transcriptase
  • a reduction or absence of a longer RT-PCR product produced using the primers spanning the region containing the included exon (e.g., including the flanking exons of the NIE) in ASO-treated cells as compared to in control ASO-treated cells indicates that splicing of the target NIE has been enhanced.
  • the exon skipping efficiency or the splicing efficiency to splice the intron containing the NIE
  • the ratio of spliced to unspliced pre-mRNA, the rate of splicing, or the extent of splicing may be improved using the ASOs described herein.
  • the amount of protein or functional RNA that is encoded by the target pre-mRNA can also be assessed to determine whether each ASO achieved the desired effect (e.g., enhanced functional protein production). Any method known in the art for assessing and/or quantifying protein production, such as Western blotting, flow cytometry, immunofluorescence microscopy, and ELISA, can be used. [0712] A second round of screening, referred to as an ASO “micro-walk” may be performed using ASOs that have been designed to hybridize to a target region of a pre-mRNA.
  • ASOs used in the ASO micro-walk are tiled every 1 nucleotide to further refine the nucleotide acid sequence of the pre- mRNA that when hybridized with an ASO results in exon skipping (or enhanced splicing of NIE).
  • Regions defined by ASOs that promote splicing of the target intron are explored in greater detail by means of an ASO “micro-walk”, involving ASOs spaced in 1-nt steps, as well as longer ASOs, typically 18-25 nt.
  • the ASO micro-walk is performed by delivering one or more ASOs, or a control ASO (an ASO with a scrambled sequence, sequence that is not expected to hybridize to the target region), for example by transfection, into a disease-relevant cell line that expresses the target pre-mRNA.
  • ASOs an ASO with a scrambled sequence, sequence that is not expected to hybridize to the target region
  • the splicing-inducing effects of each of the ASOs may be assessed by any method known in the art, for example by reverse transcriptase (RT)-PCR using primers that span the NIE, as described herein.
  • RT reverse transcriptase
  • a reduction or absence of a longer RT-PCR product produced using the primers spanning the NIE in ASO-treated cells as compared to in control ASO-treated cells indicates that exon skipping (or splicing of the target intron containing an NIE) has been enhanced.
  • the exon skipping efficiency (or the splicing efficiency to splice the intron containing the NIE), the ratio of spliced to unspliced pre-mRNA, the rate of splicing, or the extent of splicing may be improved using the ASOs described herein.
  • the amount of protein or functional RNA that is encoded by the target pre- mRNA can also be assessed to determine whether each ASO achieved the desired effect (e.g., enhanced functional protein production).
  • kits comprising: a concentrate comprising an antisense oligomer (ASO), wherein the ASO comprises a sequence with at least 80% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099; and a diluent, wherein the concentrate is miscible with the diluent; and instructions for diluting or solubilizing the ASO in the diluent.
  • ASO antisense oligomer
  • the ASO comprises a sequence with at least 80% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B. [0716] In some embodiments, the ASO comprises a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the ASO consists of a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of SEQ ID NOs: 21-67, 210-256 or 304-1099.
  • the ASO comprises a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the ASO consists of a sequence with at least 60%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 884%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.8%, 99.9%, or 100% sequence identity to any one of the sequences listed in listed in Tables 4A, 4B, 5A, 5B, 6A, 6B, 7, 8A, and 8B.
  • the diluent is an artificial cerebral spinal fluid (aCSF) solution.
  • the solution comprises a cerebral spinal fluid (CSF) sample from the subject.
  • the diluent comprises an isotonic solution.
  • the diluent comprises a phosphate-buffered (pH 6.6 – 7.6) solution.
  • the ASO as described herein is solubilized or diluted in a phosphate- buffered (pH 6.6-7.6) solution.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered (pH 6.0-8.0) solution.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered (pH 5.0-8.0) solution. In some embodiments, the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 4.5-8.5, pH 4.6-8.5, pH 4.7-8.5, pH 4.8-8.5, pH 4.9-8.5, pH5.0-8.5, pH 5.1-8.5, pH5.2-8.5, pH 5.3-8.5, pH5.4-8.5, pH 5.5-8.5, pH5.6-8.5, pH 5.7-8.5, pH 5.8-8.5, H 5.9-8.5, pH 6.0-8.5, pH 6.1-8.5, pH 6.2-8.5, pH 6.3-8.5, pH 6.4-8.5, pH 6.5-8.5, pH 6.6-8.5, pH 6.7-8.5, pH 6.8-8.5, pH 6.9-8.5, pH 7.0-8.5, pH 7.1-8.5, pH 7.2-8.5, pH 7.3-8.5, pH 7.4-8.5, pH 7.5-8.5, pH
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 4.5-8.3, pH 4.5-8.2, pH 4.5-8.1, pH 4.5- WSGR Docket No.47991-744.601 8.0, pH 4.5-7.9, pH 4.5-7.8, pH 4.5-7.7, pH 4.5-7.6, pH 4.5-7.5, pH 4.5-7.4, pH 4.5-7.3, pH 4.5-7.2, pH 4.5-7.1, pH 4.5-7.0, pH 4.5-6.9, pH 4.5-6.8, pH 4.5-6.7, pH 4.5-6.6, pH 4.5-6.5, pH 4.5-6.4, pH 4.5-6.3, pH 4.5-6.2, pH 4.5-6.1, pH 4.5-6.0, pH 4.5-5.9, pH 4.5-5.8, pH 4.5-5.7, pH 4.5-5.6, pH 4.5-5.5, pH 4.5- 5.4, pH 4.5-5.3, pH 4.5-5.2, pH 4.5-5.1, pH 4.5-5.0, pH 4.5-4.9
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-7.6, pH 6.1-7.6, pH 6.2-7.6, pH 6.3-7.6, pH 6.4-7.6, pH 6.5-7.6, pH 6.6-7.6, pH 6.7- 7.6, pH 6.8-7.6, pH 6.9-7.6, pH 7.0-7.6, pH 7.1-7.6, pH 7.2-7.6, pH 7.3-7.6, pH 7.4-7.6, or pH 7.5-7.6.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.6-8.0, pH 6.6-7.9, pH 6.6-7.8, pH 6.6-7.7, pH 6.6-7.6, pH 6.6-7.5, pH 6.6-7.4, pH 6.6- 7.3, pH 6.6-7.2, pH 6.6-7.1, pH 6.6-7.0, pH 6.6-6.9, pH 6.6-6.8, or pH 6.6-6.7.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-8.0, pH 6.1-8.0, pH 6.2-8.0, pH 6.3-8.0, pH 6.4-8.0, pH 6.5-8.0, pH 6.6-8.0, pH 6.7-8.0, pH 6.8-8.0, pH 6.9-8.0, pH 7.0-8.0, pH 7.1-8.0, pH 7.2-8.0, pH 7.3-8.0, pH 7.4-8.0, pH 7.5-8.0, pH 7.6-8.0, pH 7.7-8.0, pH 7.8- 8.0, or pH 7.9-8.0.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 6.0-7.9, pH 6.0-7.8, pH 6.0-7.7, pH 6.0-7.6, pH 6.0-7.5, pH 6.0-7.4, pH 6.0-7.3, pH 6.0-7.2, pH 6.0-7.1, pH 6.0-7.0, pH 6.0-6.9, pH 6.0-6.8, pH 6.0-6.7, pH 6.0-6.6, pH 6.0- 6.5, pH 6.0-6.4, pH 6.0-6.3, pH 6.0-6.2, or pH 6.0-6.1.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 5.7-8.5, 5.8-8.4, 5.9-8.3, 6.0-8.2, 6.1-8.1, 6.2-8.0, 6.3-7.9, 6.4-7.8, 6.5-7.7, or 6.6-7.6.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH about 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0.6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, or 8.0.
  • the ASO as described herein is solubilized or diluted in a phosphate-buffered solution with pH 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0.6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, or 8.0.
  • the ASO as described herein is solubilized or diluted in a liquid composition not buffered by a buffering agent.
  • the ASO as described herein is solubilized or diluted in a liquid composition lacks Na2HPO4 and/or NaH2PO4.
  • the liquid composition lacks phosphate ions.
  • the liquid composition comprises calcium ion, magnesium ion, and/or potassium ion.
  • the diluent comprises 25-250 mM NaCl.
  • the diluent comprises 25-250, 30-250, 35-250, 40-250, 45-250, 50-250, 55-250, 60-250, 65-250, 70-250, 75-250, 80-250, 85-250, 90-250, 95-250, 100-250, 105-250, 110-250, 115-250, 120-250, 125-250, 130-250, 135-250, 140-250, 145-250, 150-250, 155-250, 160-250, 165-250, 170-250, 175-250, 180-250, 185-250, 190-250, 195-250, 200-250, 205-250, 210-250, 215-250, 220-250, 225-250, 230-250, 235-250, 240-250, or 245-250 mM NaCl.
  • the diluent comprises 25-245, 25-240, 25-235, 25-230, 25-225, 25-220, 25-215, 25-210, 25-205, 25-200, 25-195, 25- WSGR Docket No.47991-744.601 190, 25-185, 25-180, 25-175, 25-170, 25-165, 25-160, 25-155, 25-150, 25-145, 25-140, 25-135, 25-130, 25-125, 25-120, 25-115, 25-110, 25-105, 25-110, 25-105, 25-100, 25-95, 25-90, 25-85, 25-80, 25-75, 25- 70, 25-65, 25-60, 25-55, 25-50, 25-45, 25-40, 25-35, or 25-30 mM NaCl.
  • the diluent comprises 30-245, 35-240, 40-235, 45-230, 50-225, 55-220, 60-215, 65-210, 70-205, 75-200, 80- 195, 85-190, 90-185, 95-180, 100-175, 105-170, 110-165, 115-160, 120-155, 125-150, 130-145 or 135- 140 mM NaCl.
  • the diluent comprises 100-140, 101-140, 102-140, 103-140, 104- 140, 105-140, 106-140, 107-140, 108-140, 109-140, 110-140, 111-140, 112-140, 113-140, 114-140, 115- 140, 116-140, 117-140, 118-140, 119-140, 120-140, 121-140, 122-140, 123-140, 124-140, 125-140, 126- 140, 127-140, 128-140, 129-140, 130-140, 131-140, 132-140, 133-140, 134-140, 135-140, 136-140, 137- 140, 138-140, or 139-140 mM NaCl.
  • the diluent comprises 100-139, 100-138, 100-137, 100-136, 100-135, 100-134, 100-133, 100-132, 100-131, 100-130, 100-129, 100-128, 100-127, 100-126, 100-125, 100-124, 100-123, 100-122, 100-121, 100-120, 100-119, 100-118, 100-117, 100-116, 100-115, 100-114, 100-113, 100-112, 100-111, 100-110, 100-109, 100-108, 100-107, 100-106, 100-105, 100-104, 100-103, 100-102, or 100-101 mM NaCl.
  • the diluent comprises at least 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl.
  • the diluent comprises at most 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl.
  • the diluent comprises 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, or 140 mM NaCl. [0725] In some embodiments, the diluent comprises 0.1-20 mM KCl.
  • the diluent comprises 0.1-40, 0.1-39, 0.1-38, 0.1-37, 0.1-36, 0.1-35, 0.1- 34, 0.1-33, 0.1-32, 0.1-31, 0.1-30, 0.1-29, 0.1-28, 0.1-27, 0.1-26, 0.1-25, 0.1-24, 0.1-23, 0.1-22, 0.1-21, 0.1-20, 0.1-19, 0.1-18, 0.1-17, 0.1-16, 0.1-15, 0.1-14, 0.1-13, 0.1-12, 0.1-10, 0.1-9, 0.1-8, 0.1-7, 0.1-6, 0.1-5, 0.1-4, 0.1-3, 0.1-2, or 0.1-1 mM KCl.
  • the diluent comprises 0.2-40, 0.3-40, 0.4-40, 0.5-40, 0.6-40, 0.7-40, 0.8-40, 0.9-40, 1-40, 2-40, 3-40, 4-40, 5-40, 6-40, 7-40, 8-40, 9-40, 10-40, 11-40, 12-40, 13-40, 14-40, 15-40, 16-40, 17-40, 18-40, 19-40, 20-40, 21-40, 22-40, 23-40, 24-40, 25-40, 26-40, 27-40, 28-40, 29-40, 30-40, 31-40, 32-40, 33-40, 34-40, 35-40, 36-40, 37-40, 38-40, or 39-40 mM KCl.
  • the diluent comprises 0.1-3.5, 0.2-3.5, 0.3-3.5, 0.4-3.5, 0.5-3.5, 0.6-3.5, 0.7- 3.5, 0.8-3.5, 0.9-3.5, 1.0-3.5, 1.1-3.5, 1.2-3.5, 1.3-3.5, 1.4-3.5, 1.5-3.5, 1.6-3.5, 1.7-3.5, 1.8-3.5, 1.9-3.5, 2.0-3.5, 2.1-3.5, 2.2-3.5, 2.3-3.5, 2.4-3.5, 2.5-3.5, 2.6-3.5, 2.7-3.5, 2.8-3.5, 2.9-3.5, 3.0-3.5, 3.1-3.5, 3.2- 3.5, 3.3-3.5, or 3.4-3.5 mM KCl.
  • the diluent comprises 0.1-3.4, 0.1-3.3, 0.1-3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1- 1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM KCl.
  • the diluent comprises at WSGR Docket No.47991-744.601 least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl.
  • the diluent comprises at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl.
  • the diluent comprises 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, or 3.5 mM KCl. [0727] In some embodiments, the diluent comprises 0-50 mM Na2HPO4.
  • the diluent comprises 0-50, 0-45, 0-40, 0-35, 0-30, 0-25, 0-20, 0-19, 0-18, 0-17, 0-16, 0-15, 0-14, 0-13, 0-12, 0-11, 0-10, 0-9, 0-8, 0-7, 0-6, 0-5, 0-4, 0-3, 0-2, 0-1, 0-0.9, 0-0.8, 0- 0.7, 0-0.6, 0-0.5, 0-0.4, 0-0.3, or 0-0.2 mM Na2HPO4.
  • the diluent comprises 0.1- 50, 0.1-45, 0.1-40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-19, 0.1-18, 0.1-17, 0.1-16, 0.1-15, 0.1-14, 0.1-13, 0.1-12, 0.1-11, 0.1-10, 0.1-9, 0.1-8, 0.1-7, 0.1-6, 0.1-5, 0.1-4, 0.1-3, 0.1-2, 0.1-1, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM Na 2 HPO 4 .
  • the diluent comprises 0-50, 0.1-50, 0.2-50, 0.3-50, 0.4-50, 0.5-50, 0.6-50, 0.7-50, 0.8-50, 0.9-50, 1-50, 2-50, 3-50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 1-50, 11-50, 12-50, 13-50, 14-50, 15-50, 16-50, 17-50, 18-50, 19-50, 20-50, 25- 50, 30-50, 35-50, 40-50, or 45-50 mM Na 2 HPO 4 .
  • the diluent comprises 0-20, 0.1- 20, 0.2-20, 0.3-20, 0.4-20, 0.5-20, 0.6-20, 0.7-20, 0.8-20, 0.9-20, 1-20, 2-20, 3-20, 4-20, 5-20, 6-20, 7-20, 8-20, 9-20, 10-20, 11-20, 12-20, 13-20, 14-20, 15-20, 16-20, 17-20, 18-20, or 19-20 mM Na 2 HPO 4 .
  • the diluent comprises at least 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM Na 2 HPO 4 .
  • the diluent comprises at most 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM Na 2 HPO 4 . In some embodiments, the diluent comprises 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM Na 2 HPO 4 . [0729] In some embodiments, the diluent comprises 0-50 mM NaH 2 PO 4 .
  • the diluent comprises 0-50, 0-45, 0-40, 0-35, 0-30, 0-25, 0-20, 0-19, 0-18, 0-17, 0-16, 0-15, 0-14, 0-13, 0-12, 0-11, 0-10, 0-9, 0-8, 0-7, 0-6, 0-5, 0-4, 0-3, 0-2, 0-1, 0-0.9, 0-0.8, 0- 0.7, 0-0.6, 0-0.5, 0-0.4, 0-0.3, or 0-0.2 mM NaH 2 PO 4 .
  • the diluent comprises 0.1- 50, 0.1-45, 0.1-40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-19, 0.1-18, 0.1-17, 0.1-16, 0.1-15, 0.1-14, 0.1-13, 0.1-12, 0.1-11, 0.1-10, 0.1-9, 0.1-8, 0.1-7, 0.1-6, 0.1-5, 0.1-4, 0.1-3, 0.1-2, 0.1-1, 0.1-0.9, 0.1-0.8, 0.1-0.7, 0.1-0.6, 0.1-0.5, 0.1-0.4, 0.1-0.3, or 0.1-0.2 mM NaH2PO4.
  • the diluent comprises 0-50, 0.1-50, 0.2-50, 0.3-50, 0.4-50, 0.5-50, 0.6-50, 0.7-50, 0.8-50, 0.9-50, 1-50, 2-50, 3-50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 1-50, 11-50, 12-50, 13-50, 14-50, 15-50, 16-50, 17-50, 18-50, 19-50, 20-50, 25- 50, 30-50, 35-50, 40-50, or 45-50 mM NaH2PO4.
  • the diluent comprises 0-20, 0.1- 20, 0.2-20, 0.3-20, 0.4-20, 0.5-20, 0.6-20, 0.7-20, 0.8-20, 0.9-20, 1-20, 2-20, 3-20, 4-20, 5-20, 6-20, 7-20, 8-20, 9-20, 10-20, 11-20, 12-20, 13-20, 14-20, 15-20, 16-20, 17-20, 18-20, or 19-20 mM NaH2PO4.
  • the diluent comprises at least 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM NaH2PO4.
  • the diluent comprises at most 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, WSGR Docket No.47991-744.601 16, 17, 18, 19, or 20 mM NaH2PO4. In some embodiments, the diluent comprises 0, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 mM NaH2PO4. [0731] In some embodiments, the diluent comprises 0.1-50 mM CaCl2.
  • the diluent comprises 0.1-50, 0.2-50, 0.3-50, 0.4-50, 0.5-50, 0.6-50, 0.7- 50, 0.8-50, 0.9-50, 1.0-50, -50, 1.1-50, 1.2-50, 1.3-50, 1.4-50, 1.5-50, 1.6-50, 1.7-50, 1.8-50, 1.9-50, 2.0- 50, 2.1-50, 2.2-50, 2.3-50, 2.4-50, 2.5-50, 2.6-50, 2.7-50, 2.8-50, 2.9-50, 3.0-50, 3.1-50, 3.2-50, 3.3-50, 3.4-50, 3.5-50, 3.6-50, 3.7-50, 3.8-50, 3.9-50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 10-50, 15-50, 20-50, 25- 50, 130-50, 35-50, 40-50, or 45-50 mM CaCl2.
  • the diluent comprises 0.1-45, 0.1- 40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-15, 0.1-10, 0.1-5, 0.1-4, 0.1-4.9, 0.1-4.8, 0.1-4.7, 0.1-4.6, 0.1-4.5, 0.1-4.4, 0.1-4.3, 0.1-4.2, 0.1-4.1, 0.1-4.0, 0.1-3.9, 0.1-3.8, 0.1-3.7, 0.1-3.6, 0.1-3.5, 0.1-3.4, 0.1-3.3, 0.1- 3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0,
  • the diluent comprises at least 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl 2 .
  • the diluent comprises at most 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl 2 .
  • the diluent comprises 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM CaCl 2 .
  • the diluent comprises 0.1-50 mM MgCl 2 .
  • the diluent comprises 0.1-50, 0.2-50, 0.3-50, 0.4-50, 0.5-50, 0.6-50, 0.7- 50, 0.8-50, 0.9-50, 1.0-50, -50, 1.1-50, 1.2-50, 1.3-50, 1.4-50, 1.5-50, 1.6-50, 1.7-50, 1.8-50, 1.9-50, 2.0- 50, 2.1-50, 2.2-50, 2.3-50, 2.4-50, 2.5-50, 2.6-50, 2.7-50, 2.8-50, 2.9-50, 3.0-50, 3.1-50, 3.2-50, 3.3-50, 3.4-50, 3.5-50, 3.6-50, 3.7-50, 3.8-50, 3.9-50, 4-50, 5-50, 6-50, 7-50, 8-50, 9-50, 10-50, 15-50, 20-50, 25- 50, 130-50, 35-50, 40-50, or 45-50 mM MgCl 2 .
  • the diluent comprises 0.1-45, 0.1- 40, 0.1-35, 0.1-30, 0.1-25, 0.1-20, 0.1-15, 0.1-10, 0.1-5, 0.1-4, 0.1-4.9, 0.1-4.8, 0.1-4.7, 0.1-4.6, 0.1-4.5, 0.1-4.4, 0.1-4.3, 0.1-4.2, 0.1-4.1, 0.1-4.0, 0.1-3.9, 0.1-3.8, 0.1-3.7, 0.1-3.6, 0.1-3.5, 0.1-3.4, 0.1-3.3, 0.1- 3.2, 0.1-3.1, 0.1-3.0, 0.1-2.9, 0.1-2.8, 0.1-2.7, 0.1-2.6, 0.1-2.5, 0.1-2.4, 0.1-2.3, 0.1-2.2, 0.1-2.1, 0.1-2.0, 0.1-1.9, 0.1-1.8, 0.1-1.7, 0.1-1.6, 0.1-1.5, 0.1-1.4, 0.1-1.3, 0.1-1.2, 0.1-1.1, 0.1-1.0,
  • the diluent comprises at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl2.
  • the diluent comprises at most 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl2.
  • the diluent comprises 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 mM MgCl2.
  • WSGR Docket No.47991-744.601 [0735]
  • the diluent comprises 1-100 mM NaHCO3, 1-100 mM KHCO3, or a combination thereof.
  • the diluent comprises 1-100 mM NaHCO3.
  • the diluent comprises 1-99, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM NaHCO3.
  • the diluent comprises 2-100, 3-100, 4-100, 5-100, 6-100, 7-100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60-100, 65- 100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM NaHCO3.
  • the diluent comprises 24.0-28.0, 24.0-27.9, 24.0-27.8, 24.0-27.7, 24.0-27.6, 24.0-27.5, 24.0-27.4, 24.0-27.3, 24.0-27.2, 24.0-27.1, 24.0-27.0, 24.0-26.9, 24.0-26.8, 24.0-26.7, 24.0-26.6, 24.0-26.5, 24.0-26.4, 24.0- 26.3, 24.0-26.2, 24.0-26.1, 24.0-26.0, 24.0-25.9, 4.0-25.8, 24.0-25.7, 24.0-25.6, 24.0-25.5, 24.0-25.4, 24.0-25.3, 24.0-25.2, 24.0-25.1, 24.0-25.0, 24.0-24.9, 24.0-24.8, 24.0-24.7, 24.0-24.6, 24.0-24.5, 24.0- 24.4, 24.0-24.3, 24.0-24.2, or 24.0-24.1 mM NaHCO 3 .
  • the diluent comprises 24.1- 28.0, 24.2-28.0, 24.3-28.0, 24.4-28.0, 24.5-28.0, 24.6-28.0, 24.7-28.0, 24.8-28.0, 24.9-28.0, 25.0-28.0, 25.1-28.0, 25.2-28.0, 25.3-28.0, 25.4-28.0, 25.5-28.0, 25.6-28.0, 25.7-28.0, 25.8-28.0, 25.9-28.0, 26.0- 28.0, 26.1-28.0, 26.2-28.0, 26.3-28.0, 26.4-28.0, 26.5-28.0, 26.6-28.0, 26.7-28.0, 26.8-28.0, 26.9-28.0, 27.0-28.0, 27.1-28.0, 27.2-28.0, 27.3-28.0, 27.4-28.0, 27.5-28.0, 27.6-28.0, 27.7-28.0, 27.8-28.0, or 27.9- 28.0 mM NaHCO 3 .
  • the diluent comprises at least 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.0, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO 3 .
  • the diluent comprises at most 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO 3 .
  • the diluent comprises 24.0, 24.1, 24.2, 24.3, 24.4, 24.5, 24.6, 24.7, 24.8, 24.9, 25.0, 25.1, 25.2, 25.3, 25.4, 25.5, 25.6, 25.7, 25.8, 25.9, 26.1, 26.2, 26.3, 26.4, 26.5, 26.6, 26.7, 26.8, 26.9, 27.0, 27.1, 27.2, 27.3, 27.4, 27.5, 27.6, 27.7, 27.8, 27.9, or 28.0 mM NaHCO 3 .
  • the diluent comprises 1-100 mM KHCO 3 .
  • the diluent comprises 1-99, 1-95, 1-90, 1-85, 1-80, 1-75, 1-70, 1-65, 1-60, 1-55, 1-50, 1-45, 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, or 1-2 mM KHCO3.
  • the diluent comprises 2-100, 3-100, 4-100, 5-100, 6-100, 7-100, 8-100, 9-100, 10-100, 15-100, 20-100, 25-100, 30-100, 35-100, 40-100, 45-100, 50-100, 55-100, 60-100, 65- 100, 70-100, 75-100, 80-100, 85-100, 90-100, or 95-100 mM KHCO3.
  • the diluent comprises 24.0-28.0, 24.0-27.9, 24.0-27.8, 24.0-27.7, 24.0-27.6, 24.0-27.5, 24.0-27.4, 24.0-27.3, 24.0- 27.2, 24.0-27.1, 24.0-27.0, 24.0-26.9, 24.0-26.8, 24.0-26.7, 24.0-26.6, 24.0-26.5, 24.0-26.4, 24.0-26.3, 24.0-26.2, 24.0-26.1, 24.0-26.0, 24.0-25.9, 4.0-25.8, 24.0-25.7, 24.0-25.6, 24.0-25.5, 24.0-25.4, 24.0- 25.3, 24.0-25.2, 24.0-25.1, 24.0-25.0, 24.0-24.9, 24.0-24.8, 24.0-24.7, 24.0-24.6, 24.0-24.5, 24.0-24.4, 24.0-24.3, 24.0-24.2, or 24.0-24.1 mM KHCO3.
  • the diluent comprises 24.1-28.0, 24.2-28.0, 24.3-28.0, 24.4-28.0, 24.5-28.0, 24.6-28.0, 24.7-28.0, 24.8-28.0, 24.9-28.0, 25.0-28.0, 25.1- WSGR Docket No.47991-744.601 28.0, 25.2-28.0, 25.3-28.0, 25.4-28.0, 25.5-28.0, 25.6-28.0, 25.7-28.0, 25.8-28.0, 25.9-28.0, 26.0-28.0, 26.1-28.0, 26.2-28.0, 26.3-28.0, 26.4-28.0, 26.5-28.0, 26.6-28.0, 26.7-28.0, 26.8-28.0, 26.9-28.0, 27.0- 28.0, 27.1-28.0, 27.2-28.0, 27.3-28.0, 27.4-28.0, 27.5-28.0, 27.6-28.0, 27.7-28.0, 27.8-28.0, or 27.9-28.0 mM KHCO3.

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Abstract

L'invention concerne une formulation pharmaceutique comprenant un oligomère antisens (ASO), un kit et une méthode d'utilisation de celle-ci pour traiter une maladie ou un état pathologique par modulation de l'expression de la protéine NaV1.1 dans les cellules d'un sujet en ayant besoin, l'ASO modulant l'épissage d'un exon induisant une décroissance d'ARNm à médiation non-sens à partir d'un pré-ARNm dans les cellules du sujet, modulant ainsi l'expression de la protéine NaV1.1 chez le sujet.
PCT/US2024/015838 2023-02-14 2024-02-14 Formulations d'oligomères antisens WO2024173582A2 (fr)

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US20030087861A1 (en) * 2001-05-17 2003-05-08 Iversen Patrick L Combined approach to treatment of cancer using a c-myc antisense oligomer
BR112020026542A2 (pt) * 2018-06-26 2021-04-06 Nippon Shinyaku Co., Ltd. Composição que compreende oligonucleotídeo antissenso e uso da mesma para tratamento de distrofia muscular de duchenne
JP2023504195A (ja) * 2019-12-06 2023-02-01 ストーク セラピューティクス，インク． 状態および疾患の処置のためのアンチセンスオリゴマー

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