WO2023028575A2 - Compounds and methods for modulating scn1a expression - Google Patents

Compounds and methods for modulating scn1a expression Download PDF

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WO2023028575A2
WO2023028575A2 PCT/US2022/075493 US2022075493W WO2023028575A2 WO 2023028575 A2 WO2023028575 A2 WO 2023028575A2 US 2022075493 W US2022075493 W US 2022075493W WO 2023028575 A2 WO2023028575 A2 WO 2023028575A2
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seq
oligomeric compound
modified
certain embodiments
intemucleoside linkage
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WO2023028575A3 (en
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Paymaan JAFAR-NEJAD
Frank Rigo
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Ionis Pharmaceuticals Inc
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Ionis Pharmaceuticals Inc
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Priority to JP2024513011A priority Critical patent/JP2024534157A/ja
Priority to EP22862285.8A priority patent/EP4392562A4/en
Priority to US18/685,362 priority patent/US20240360449A1/en
Priority to AU2022334739A priority patent/AU2022334739A1/en
Priority to CA3230299A priority patent/CA3230299A1/en
Priority to IL310811A priority patent/IL310811A/en
Priority to MX2024002553A priority patent/MX2024002553A/es
Priority to CN202280065211.6A priority patent/CN118451183A/zh
Priority to KR1020247009728A priority patent/KR20240049349A/ko
Publication of WO2023028575A2 publication Critical patent/WO2023028575A2/en
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Definitions

  • oligomeric compounds, methods, and pharmaceutical compositions for modulating expression of SCN1A RNA and/or protein in a cell or subject are useful to ameliorate at least one symptom of a developmental or epileptic encephalopathic disease, such as, for example, Dravet Syndrome.
  • symptoms include seizures, sudden unexpected death in epilepsy, status epilepticus, behavioral dysfunctions, movement and balance dysfunctions, orthopedic conditions, motor dysfunctions, cognitive impairment, delayed language and speech, visual motor integration dysfunctions, visual perception dysfunctions, executive dysfunctions, and dysautonomia.
  • the human gene SCN1A encodes human SCN1A protein, the alpha-1 subunit of the voltage-gated sodium channel NaVl.1. Mutations in SCN1A lead to developmental and epileptic encephalopathies (DEEs), including Dravet Syndrome (previously known as Severe Myoclonic Epilepsy of Infancy (SMEI)), one of the most severe childhood forms of epilepsy; other epileptic disorders, including, for example, Genetic Epilepsy with Febrile Seizures Plus (GEFS+) and other febrile seizures, Idiopathic/Generic Generalized Epilepsies (IGE/GGE), Temporal Lobe Epilepsy, Myoclonic Astatic Epilepsy (MAE), Lennox-Gastaut Syndrome, and Migrating Partial Epilepsy of Infancy (MMPSI); and familial hemiplegic migraines, with or without epilepsy (Harkin, L.A., et al., 2007, Brain 130, 843-852; Escayg
  • DEEs are associated with SCN1A haploinsufficiency (Parihar, R., et al., 2013, J. Human Genetics, 58, 573- 580).
  • Symptoms associated with DEEs include seizures that are prolonged in duration (often lasting longer than 10 minutes), frequent seizures (for example, convulsive, myoclonic, absence, focal, obtundation status, and tonic seizures), sudden unexpected death in epilepsy, status epilepticus, behavioral dysfunctions (for example, aggressiveness, agitation, obsessiveness, preservation, hoarding behavior, or sleep disorders), movement and balance dysfunctions, orthopedic conditions, motor system dysfunctions (for example, ataxia, tremors, dysarthria, pyramidal, and extrapyramidal signs), cognitive impairment, delayed language and speech, visual motor integration dysfunctions, visual perception dysfunctions, executive dysfunctions, and dysautonomia. Dravet Syndrome patients experience additional neurodevelopmental delays, leading to severe neurological disability (Guzzetta, F.
  • NIE decay-inducing exon
  • the amount of SCN1A RNA and/or SCN1A protein is increased.
  • the compounds, methods, or pharmaceutical compositions modulate splicing of SCN1A RNA.
  • the amount of full-length SCN1A RNA and/or full-length SCN1A protein is increased.
  • the amount of SCN1A RNA including an NIE is reduced.
  • the amount of SCN1A RNA excluding an NIE is increased.
  • the NIE is NIE-1.
  • the compounds, methods, and pharmaceutical compositions are useful for treating a disease or disorder associated with SCN1A.
  • the disease or disorder associated with SCN1A is an SCN1A haploinsufficiency.
  • the disease or disorder associated with SCN1A is a developmental or epileptic encephalopathic disease (DEE).
  • the developmental or epileptic encephalopathic disease is any of Genetic Epilepsy with Febrile Seizures Plus (GEFS+), febrile seizures, Idiopathic/Generic Generalized Epilepsies (IGE/GGE), Temporal Lobe Epilepsy, Myoclonic Astatic Epilepsy (MAE), Lennox-Gastaut Syndrome, or Migrating Partial Epilepsy of Infancy (MMPSI).
  • the developmental or epileptic encephalopathic disease is Dravet Syndrome.
  • the DEE is treated by increasing the amount of full-length SCN1A RNA and/or full-length SCN1A protein in a subject, or cell thereof, with compounds capable of excluding an NIE from an SCN1A RNA.
  • exclusion of an NIE from an SCN1A RNA reduces or prevents degradation of the SCN1A transcript via the NMD pathway.
  • exclusion of an NIE from an SCN1A RNA increases full-length SCN1A RNA and/or full-length SCN1A protein wherein removal of the NIE prevents degradation of the SCN1A transcript via the NMD pathway.
  • compounds useful for modulating splicing of SCN1A RNA are oligomeric compounds.
  • the oligomeric compound comprises or consists of a modified oligonucleotide.
  • the disease or disorder associated with SCN1A is a DEE.
  • the DEE is Dravet Syndrome.
  • symptoms or hallmarks of the DEE include seizures that are prolonged in duration (often lasting longer than 10 minutes), frequent seizures (for example, convulsive, myoclonic, absence, focal, obtundation status, and tonic seizures), sudden unexpected death in epilepsy, status epilepticus, behavioral dysfunctions (for example, aggressiveness, agitation, obsessiveness, preservation, hoarding behavior, or sleep disorders), movement and balance dysfunctions, orthopedic conditions, motor system dysfunctions (for example, ataxia, tremors, dysarthria, pyramidal, and extrapyramidal signs), cognitive impairment, delayed language and speech, visual motor integration dysfunctions, visual perception dysfunctions, executive dysfunctions, and dysautonomia Detailed Description
  • 2’-deoxynucleoside means a nucleoside comprising a 2’-H(H) deoxyfuranosyl sugar moiety.
  • a 2’-deoxynucleoside is a 2’-p-D-deoxynucleoside and comprises a 2’-p-D-deoxyribosyl sugar moiety, which has the P-D ribosyl configuration as found in naturally occurring deoxyribonucleic acids (DNA).
  • a 2’-deoxynucleoside may comprise a modified nucleobase or may comprise an RNA nucleobase (uracil).
  • 2 ’-MOE means a 2’-O(CH 2 )2OCH 3 group in place of the 2 ’-OH group of a furanosyl sugar moiety.
  • a “2 ’-MOE sugar moiety” or a “2’-O-methoxyethyl sugar moiety” means a sugar moiety with a 2’- OCH2CH2OCH 3 group in place of the 2’-OH group of a furanosyl sugar moiety. Unless otherwise indicated, a 2’-MOE sugar moiety is in the P-D-ribosyl configuration. “MOE” means O-methoxyethyl.
  • 2’-MOE nucleoside means a nucleoside comprising a 2’-MOE sugar moiety.
  • NMA means O-N-methyl acetamide.
  • 2’-NMA nucleoside means a nucleoside comprising a 2’-NMA sugar moiety.
  • 2 ’-substituted nucleoside means a nucleoside comprising a 2’-substituted furanosyl sugar moiety.
  • 2 ’-substituted in reference to a sugar moiety means a sugar moiety comprising at least one 2'- substituent group other than H or OH.
  • 5-methylcytosine means a cytosine modified with a methyl group attached to the 5 position. A 5-methylcytosine is a modified nucleobase.
  • administering means providing a pharmaceutical agent to a subject.
  • “ameliorate” in reference to a treatment means improvement in at least one symptom or hallmark relative to the same symptom or hallmark in the absence of the treatment.
  • amelioration is the reduction in the severity or frequency of a symptom or hallmark or the delayed onset or slowing of progression in the severity or frequency of a symptom or hallmark.
  • the symptom or hallmark is seizures that are prolonged in duration (often lasting longer than 10 minutes), frequent seizures (for example, convulsive, myoclonic, absence, focal, obtundation status, and tonic seizures), sudden unexpected death in epilepsy, status epilepticus, behavioral dysfunctions (for example, aggressiveness, agitation, obsessiveness, preservation, hoarding behavior, or sleep disorders), and developmental delays, movement and balance dysfunctions, orthopedic conditions, motor system and cognitive dysfunctions (for example, ataxia, tremors, dysarthria, pyramidal, and extrapyramidal signs), cognitive impairment, delayed language and speech issues, visual motor integration dysfunctions, visual perception dysfunctions, executive dysfunctions, growth and nutrition issues, sleeping difficulties, chronic infections, sensory integration disorders, or dysautonomia.
  • seizures that are prolonged in duration (often lasting longer than 10 minutes), frequent seizures (for example, convulsive, myoclonic, absence, focal, obtundation status, and tonic seizures), sudden unexpected death in epilepsy, status epi
  • Cerebrospinal fluid or “CSF” means the fluid filling the space around the brain and spinal cord.
  • Artificial cerebrospinal fluid” or “aCSF” means a prepared or manufactured fluid that has certain properties (e.g., osmolarity, pH, and/or electrolytes) of cerebrospinal fluid and is biocompatible with CSF.
  • conjugate group means a group of atoms that is directly attached to an oligonucleotide.
  • Conjugate groups include a conjugate moiety and a conjugate linker that attaches the conjugate moiety to the oligonucleotide.
  • conjugate linker means a single bond or a group of atoms comprising at least one bond that connects a conjugate moiety to an oligonucleotide.
  • conjugate moiety means a group of atoms that modifies one or more properties of a molecule compared to the identical molecule lacking the conjugate moiety, including but not limited to pharmacodynamics, pharmacokinetics, stability, binding, absorption, tissue distribution, cellular distribution, cellular uptake, charge and clearance.
  • intemucleoside linkage is the covalent linkage between adjacent nucleosides in an oligonucleotide.
  • modified intemucleoside linkage means any intemucleoside linkage other than a phosphodiester intemucleoside linkage.
  • linked nucleosides are nucleosides that are connected in a contiguous sequence (i.e., no additional nucleosides are presented between those that are linked).
  • linker-nucleoside means a nucleoside that links, either directly or indirectly, an oligonucleotide to a conjugate moiety. Linker-nucleosides are located within the conjugate linker of an oligomeric compound. Linker- nucleosides are not considered part of the oligonucleotide portion of an oligomeric compound even if they are contiguous with the oligonucleotide.
  • motif means the pattern of unmodified and/or modified sugar moieties, nucleobases, and/or intemucleoside linkages, in an oligonucleotide.
  • nonsense-mediated decay -inducing exon NDE is an exon, or a pseudo-exon, that, when included in an mRNA transcript can activate the nonsense-mediated decay (NMD) pathway.
  • NIE-1 is a 64 nucleobase in length NIE located in intron 20 of the SCN1A gene (chr2: 166863579-166864271, hgl9; Carvill et al., 2018), which, when present in the transcript, causes degradation of the SCN1A transcript.
  • human NIE-1 has the nucleobase sequence of SEQ ID NO: 16.
  • mouse NIE-1 has the nucleobase sequence of SEQ ID NO: 17.
  • modified nucleoside means a nucleoside comprising a modified nucleobase and/or a modified sugar moiety.
  • nucleobase means an unmodified nucleobase or a modified nucleobase.
  • a nucleobase is a heterocyclic moiety.
  • an “unmodified nucleobase” is adenine (A), thymine (T), cytosine (C), uracil (U), or guanine (G).
  • a “modified nucleobase” is a group of atoms other than unmodified A, T, C, U, or G capable of pairing with at least one other nucleobase.
  • a “5-methylcytosine” is a modified nucleobase.
  • a universal base is a modified nucleobase that can pair with any one of the five unmodified nucleobases.
  • nucleobase sequence means the order of contiguous nucleobases in a nucleic acid or oligonucleotide independent of any sugar or intemucleoside linkage modification.
  • nucleoside means a compound or fragment of a compound comprising a nucleobase and a sugar moiety.
  • the nucleobase and sugar moiety are each, independently, unmodified or modified.
  • oligomeric compound means an oligonucleotide and optionally one or more additional features, such as a conjugate group or terminal group.
  • An oligomeric compound may be paired with a second oligomeric compound that is complementary to the first oligomeric compound or may be unpaired.
  • a “singled-stranded oligomeric compound” is an unpaired oligomeric compound.
  • oligomeric duplex means a duplex formed by two oligomeric compounds having complementary nucleobase sequences.
  • oligonucleotide means a strand of linked nucleosides connected via intemucleoside linkages, wherein each nucleoside and intemucleoside linkage may be modified or unmodified. Unless otherwise indicated, oligonucleotides consist of 8-50 linked nucleosides.
  • modified oligonucleotide means an oligonucleotide, wherein at least one nucleoside or intemucleoside linkage is modified.
  • unmodified oligonucleotide means an oligonucleotide that does not comprise any nucleoside modifications or intemucleoside modifications.
  • pharmaceutically acceptable carrier or diluent means any substance suitable for use in administering to an animal. Certain such carriers enable pharmaceutical compositions to be formulated as, for example, tablets, pills, dragees, capsules, liquids, gels, symps, slurries, suspension, and lozenges for the oral ingestion by a subject.
  • a pharmaceutically acceptable diluent is sterile water, sterile saline, sterile buffer solution or sterile artificial cerebrospinal fluid.
  • pharmaceutically acceptable salts means physiologically and pharmaceutically acceptable salts of compounds. Pharmaceutically acceptable salts retain the desired biological activity of the parent compound and do not impart undesired toxicological effects thereto.
  • pharmaceutical composition means a mixture of substances suitable for administering to a subject.
  • a pharmaceutical composition may comprise an oligomeric compound and a sterile aqueous solution.
  • a pharmaceutical composition shows activity in free uptake assay in certain cell lines.
  • stereorandom or “stereorandom chiral center” in the context of a population of molecules of identical molecular formula means a chiral center that is not controlled during synthesis, or enriched following synthesis, for a particular absolute stereochemical configuration.
  • the stereochemical configuration of a chiral center is random when it is the result of a synthetic method that is not designed to control the stereochemical configuration.
  • the number of molecules having the (S) configuration of the stereorandom chiral center may be but is not necessarily the same as the number of molecules having the (R) configuration of the stereorandom chiral center (“racemic”).
  • the stereorandom chiral center is not racemic because one absolute configuration predominates following synthesis, e.g., due to the action of non-chiral reagents near the enriched stereochemistry of an adjacent sugar moiety.
  • a stereorandom chiral center is a stereorandom phosphorothioate intemucleoside linkage.
  • subject means a human or non-human animal.
  • sugar moiety means an unmodified sugar moiety or a modified sugar moiety.
  • unmodified sugar moiety means a 2’-OH(H) ribosyl moiety, as found in RNA (an “unmodified RNA sugar moiety”), or a 2’-H(H) deoxyribosyl sugar moiety, as found in DNA (an “unmodified DNA sugar moiety”).
  • Unmodified sugar moieties have one hydrogen at each of the 1 ’, 3 ’, and 4’ positions, an oxygen at the 3 ’ position, and two hydrogens at the 5’ position.
  • modified sugar moiety or “modified sugar” means a modified furanosyl sugar moiety or a sugar surrogate.
  • sugar surrogate means a modified sugar moiety that can link a nucleobase to another group, such as an intemucleoside linkage, conjugate group, or terminal group in an oligonucleotide, but which is not a furanosyl sugar moiety or a bicyclic sugar moiety.
  • Modified nucleosides comprising sugar surrogates can be incorporated into one or more positions within an oligonucleotide and such oligonucleotides are capable of hybridizing to complementary oligomeric compounds or target nucleic acids.
  • sugar surrogates include GNA (glycol nucleic acid), FHNA (fluoro hexitol nucleic acid), morpholino, and other structures described herein and known in the art.
  • symptom or hallmark means any physical feature or test result that indicates the existence or extent of a disease or disorder.
  • a symptom is apparent to a subject or to a medical professional examining or testing said subject.
  • a hallmark is apparent upon invasive diagnostic testing, including, but not limited to, post-mortem tests.
  • a hallmark is apparent on a brain MRI scan.
  • target nucleic acid and “target RNA” mean a nucleic acid that an oligomeric compound is designed to affect.
  • Target RNA means an RNA transcript and includes pre-mRNA and mRNA unless otherwise specified.
  • target region means a portion of a target nucleic acid to which an oligomeric compound is designed to hybridize.
  • terminal group means a chemical group or group of atoms that is covalently linked to a terminus of an oligonucleotide.
  • antisense activity means any detectable and/or measurable change attributable to the hybridization of an antisense compound to its target nucleic acid. In certain embodiments, antisense activity is a decrease in the amount or expression of a target nucleic acid or protein encoded by such target nucleic acid compared to target nucleic acid levels or target protein levels in the absence of the antisense compound. In certain embodiments, antisense activity is the modulation of splicing of a target pre-mRNA.
  • antisense agent means an antisense compound and optionally one or more additional features, such as a sense compound.
  • antisense compound means an antisense oligonucleotide and optionally one or more additional features, such as a conjugate group.
  • sense compound means a sense oligonucleotide and optionally one or more additional features, such as a conjugate group.
  • antisense oligonucleotide means an oligonucleotide, including the oligonucleotide portion of an antisense compound, that is capable of hybridizing to a target nucleic acid and is capable of at least one antisense activity.
  • Antisense oligonucleotides include but are not limited to antisense RNAi oligonucleotides and antisense RNase H oligonucleotides.
  • sense oligonucleotide means an oligonucleotide, including the oligonucleotide portion of a sense compound, that is capable of hybridizing to an antisense oligonucleotide.
  • hybridization means the annealing of oligonucleotides and/or nucleic acids. While not limited to a particular mechanism, the most common mechanism of hybridization involves hydrogen bonding, which may be Watson-Crick, Hoogsteen or reversed Hoogsteen hydrogen bonding, between complementary nucleobases.
  • complementary nucleic acid molecules include, but are not limited to, an antisense compound and a nucleic acid target. In certain embodiments, complementary nucleic acid molecules include, but are not limited to, an oligonucleotide and a nucleic acid target.
  • treating means improving a subject’s disease or condition by administering an oligomeric compound described herein.
  • treating a subject improves a symptom relative to the same symptom in the absence of the treatment.
  • treatment reduces in the severity or frequency of a symptom, or delays the onset of a symptom, slows the progression of a symptom, or slows the severity or frequency of a symptom.
  • terapéuticaally effective amount means an amount of a pharmaceutical agent or composition that provides a therapeutic benefit to a subject. For example, a therapeutically effective amount improves a symptom of a disease.
  • Embodiment 1 An oligomeric compound comprising a modified oligonucleotide according to the following chemical notation:
  • A an adenine nucleobase
  • mC a 5-methylcytosine nucleobase
  • G a guanine nucleobase
  • T a thymine nucleobase
  • n a 2’-NMA sugar moiety
  • s a phosphorothioate intemucleoside linkage
  • o a phosphodiester intemucleoside linkage.
  • Embodiment 2 The oligomeric compound of embodiment 1, consisting of the modified oligonucleotide.
  • Embodiment 3 The oligomeric compound of embodiment 1 or embodiment 2, wherein the modified oligonucleotide is a free acid.
  • Embodiment 4 The oligomeric compound of embodiment 1 or embodiment 2, wherein the modified oligonucleotide is a salt.
  • Embodiment 5 The oligomeric compound of embodiment 4, wherein the modified oligonucleotide is a sodium salt or a potassium salt.
  • Embodiment 6 An oligomeric compound comprising a modified oligonucleotide consisting of 17 to 30 linked nucleosides and having a nucleobase sequence comprising at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, or at least 25 consecutive nucleobases of any of the nucleobase sequences of SEQ ID NOs: 19-22 or 63-86, wherein the modified oligonucleotide comprises at least one modification selected from a modified sugar moiety and a modified intemucleoside linkage.
  • Embodiment 7 The oligomeric compound of embodiment 6, wherein the modified oligonucleotide consists of 18-25 linked nucleosides.
  • Embodiment 8 The oligomeric compound of any embodiments 6-7, wherein the modified oligonucleotide consists of 18, 23 or 25 linked nucleosides.
  • Embodiment 9 The oligomeric compound according to embodiment 6, wherein the nucleobase sequence of the modified oligonucleotide comprises the nucleobase sequence of any of SEQ ID Nos: 19-22 or 63-86.
  • Embodiment 10 The oligomeric compound according to embodiment 6, wherein the nucleobase sequence of the modified oligonucleotide consists of the nucleobase sequence of any of SEQ ID Nos: 19-22 or 63-86.
  • Embodiment 11 The oligomeric compound according to any of embodiments 6-10, wherein the modified oligonucleotide comprises at least one modified sugar moiety.
  • Embodiment 12 The oligomeric compound of any of embodiment 11, wherein the modified oligonucleotide comprises at least one non-bicyclic modified sugar moiety.
  • Embodiment 13 The oligomeric compound of embodiment 12, wherein the non-bicyclic modified sugar moiety is a 2 ’-MOE sugar moiety or a 2’-NMA sugar moiety.
  • Embodiment 14 The oligomeric compound of any of embodiments 11-13, wherein each nucleoside of the modified oligonucleotide comprises a modified sugar moiety.
  • Embodiment 15 The oligomeric compound of any of embodiments 11-14, wherein each modified sugar moiety is a 2’-NMA sugar moiety.
  • Embodiment 16 The oligomeric compound of any of embodiments 6-15, wherein the modified oligonucleotide comprises at least one modified intemucleoside linkage.
  • Embodiment 17 The oligomeric compound of embodiment 16, wherein the at least one modified intemucleoside linkage is a phosphorothioate intemucleoside linkage.
  • Embodiment 18 The oligomeric compound of embodiment 16 or embodiment 17, wherein the modified oligonucleotide comprises at least one phosphodiester intemucleoside linkage.
  • Embodiment 19 The oligomeric compound of any of embodiments 16-18, wherein each intemucleoside linkage is independently selected from a phosphodiester intemucleoside linkage and a phosphorothioate intemucleoside linkage.
  • Embodiment 20 The oligomeric compound of any of embodiments 16, 17, or 19, wherein each intemucleoside linkage is a phosphorothioate intemucleoside linkage.
  • Embodiment 21 The oligomeric compound of any of embodiments 6-20, wherein the modified oligonucleotide comprises at least one modified nucleobase.
  • Embodiment 22 The oligomeric compound of embodiment 21, wherein the modified nucleobase is a 5 -methyl cytosine
  • Embodiment 23 An oligomeric compound comprising a modified oligonucleotide according to the following chemical notation: (SEQ ID NO: 45)' (SEQ ID NO: 46)'
  • A an adenine nucleobase
  • mC a 5-methylcytosine nucleobase
  • G a guanine nucleobase
  • T a thymine nucleobase
  • n a 2’-NMA sugar moiety
  • s a phosphorothioate intemucleoside linkage
  • o a phosphodiester linkage.
  • Embodiment 24 The oligomeric compound of embodiment 23, consisting of the modified oligonucleotide.
  • Embodiment 25 The oligomeric compound of embodiment 23 or embodiment 24, wherein the modified oligonucleotide is a free acid.
  • Embodiment 26 The oligomeric compound of embodiment 23 or embodiment 24, wherein the modified oligonucleotide is a salt.
  • Embodiment 27 The oligomeric compound of embodiment 26, wherein the modified oligonucleotide is a sodium salt or a potassium salt.
  • Embodiment 28 A population of oligomeric compounds of any of embodiments 1-27, wherein all of the phosphorothioate intemucleoside linkages of the modified oligonucleotide are stereorandom.
  • Embodiment 29 A pharmaceutical composition comprising an oligomeric compound of any of embodiments 1-27 or a population of oligomeric compounds of embodiment 28, and a pharmaceutically acceptable diluent.
  • Embodiment 30 The pharmaceutical composition of embodiment 29, wherein the pharmaceutically acceptable diluent is artificial cerebrospinal fluid (aCSF) or PBS.
  • aCSF artificial cerebrospinal fluid
  • Embodiment 31 The pharmaceutical composition of embodiment 29 or embodiment 30, wherein the pharmaceutical composition consists essentially of the oligomeric compound and aCSF or PBS.
  • Embodiment 32 The pharmaceutical composition of any of embodiments 29-31, wherein the pharmaceutical composition consists essentially of the population of modified oligonucleotides or the population of oligomeric compounds and aCSF or PBS.
  • Embodiment 33 A method comprising administering to a subject an oligomeric compound of any of embodiments 1-27, a population of oligomeric compounds of embodiment 28, or a pharmaceutical composition of any of embodiments 29-32.
  • Embodiment 34 A method of treating a disease associated with SCN1A comprising administering to a subject having a disease associated with SCN1A a therapeutically effective amount of an oligomeric compound of any of embodiments 1-27, a population of oligomeric compounds of embodiment 28, or a pharmaceutical composition of any of embodiments 29-32, thereby treating the disease associated with SCN1A.
  • Embodiment 35 The method of embodiment 34, wherein the disease associated with SCN1A is a developmental or epileptic encephalopathic disease.
  • Embodiment 36 The method of embodiment 35, wherein the developmental or epileptic encephalopathic disease is Dravet Syndrome.
  • Embodiment 37 The method of embodiment 35 or embodiment 36, wherein the developmental or epileptic encephalopathic disease is any of Genetic Epilepsy with Febrile Seizures Plus (GEFS+), febrile seizures, Idiopathic/Generic Generalized Epilepsies (IGE/GGE), Temporal Lobe Epilepsy, Myoclonic Astatic Epilepsy (MAE), Lennox-Gastaut Syndrome, or Migrating Partial Epilepsy of Infancy (MMPSI).
  • GEFS+ Genetic Epilepsy with Febrile Seizures Plus
  • IGE/GGE Idiopathic/Generic Generalized Epilepsies
  • MAE Myoclonic Astatic Epilepsy
  • Lennox-Gastaut Syndrome or Migrating Partial Epilepsy of Infancy
  • Embodiment 38 The method of any of embodiments 33-37, wherein administering the oligomeric compound, the population of oligomeric compounds, or the pharmaceutical composition reduces the frequency of seizures, reduces the duration of seizures, reduces status epilepticus, improves behavioral functions, improves movement and balance, improves orthopedic conditions, improves motor functions, reduces cognitive impairment, improves language and speech, improves visual motor integration functions, improvise visual perception functions, improves executive functions, or reduces dysautonomia.
  • Embodiment 39 The method of embodiment 38, wherein the seizures are frequent or prolonged in duration.
  • Embodiment 40 The method of embodiment 38 or embodiment 39, wherein the seizure is any of convulsive, myoclonic, absence, focal, obtundation status, or tonic.
  • Embodiment 41 The method of any of embodiments 33-40, wherein the frequency of seizures is reduced.
  • Embodiment 42 The method of any of embodiments 33-41, wherein the duration of seizures is reduced.
  • Embodiment 43 The method of any of embodiments 33-42, wherein the subject is human.
  • Embodiment 44 A method of increasing expression of SCN1A in a cell comprising contacting the cell with an oligomeric compound of any of embodiments 1-27, a population of oligomeric compounds of embodiment 28, or a pharmaceutical composition of any of embodiments 29-32.
  • Embodiment 45 A method of modulating splicing of an SCN1A RNA in a cell comprising contacting the cell with an oligomeric compound of any of embodiments 1-27.
  • Embodiment 46 The method of embodiment 45, wherein the amount of SCN1A RNA that includes an NIE is reduced.
  • Embodiment 47 The method of embodiment 45 or embodiment 46, wherein the amount of SCN1A RNA that includes NIE-1 is reduced.
  • Embodiment 48 The method of any of embodiments 45-47, wherein the amount of SCN1A RNA that excludes an NIE is increased.
  • Embodiment 49 The method of any of embodiments 45-48, wherein the amount of SCN1A RNA that excludes NIE-1 is increased.
  • Embodiment 50 The method of any of embodiments 45-49, wherein the cell is a cerebral cortex, hippocampus, brainstem, or thalamus cell.
  • Embodiment 51 The method of any of embodiments 45-50, wherein the cell is a human cell.
  • Embodiment 52 Use of an oligomeric compound of any of embodiments 1-27, a population of oligomeric compounds of embodiment 28, or a pharmaceutical composition of any of embodiments 29-32 for treating a disease associated with SCN1A.
  • Embodiment 53 Use of an oligomeric compound of any of embodiments 1-27 a population of oligomeric compounds of embodiment 28, or a pharmaceutical composition of any of embodiments 29-32 in the manufacture of a medicament for treating a disease associated with SCN1A.
  • Embodiment 54 The use of embodiment 52 or embodiment 53, wherein the disease associated with SCN1A is a developmental or epileptic encephalopathic disease.
  • Embodiment 55 The use of embodiment 54, wherein the developmental or epileptic encephalopathic disease is Dravet Syndrome.
  • Embodiment 56 The use of embodiment 54 or embodiment 55, wherein the developmental or epileptic encephalopathic disease is any of Genetic Epilepsy with Febrile Seizures Plus (GEFS+), febrile seizures, Idiopathic/Generic Generalized Epilepsies (IGE/GGE), Temporal Lobe Epilepsy, Myoclonic Astatic Epilepsy (MAE), Lennox-Gastaut Syndrome, or Migrating Partial Epilepsy of Infancy (MMPSI).
  • GEFS+ Genetic Epilepsy with Febrile Seizures Plus
  • IGE/GGE Idiopathic/Generic Generalized Epilepsies
  • MAE Myoclonic Astatic Epilepsy
  • Lennox-Gastaut Syndrome or Migrating Partial Epilepsy of Infancy (MMPSI).
  • compounds are represented by the chemical notations in the following table.
  • compounds are represented by the following chemical notations (5’ to 3’): wherein,
  • A an adenine nucleobase
  • mC a 5-methylcytosine nucleobase
  • G a guanine nucleobase
  • T a thymine nucleobase
  • n a 2’-NMA sugar moiety
  • s a phosphorothioate intemucleoside linkage
  • o a phosphodiester intemucleoside linkage.
  • oligomeric compounds comprising oligonucleotides, which consist of linked nucleosides.
  • Oligonucleotides may be unmodified oligonucleotides (RNA or DNA) or may be modified oligonucleotides.
  • Modified oligonucleotides comprise at least one modification relative to unmodified RNA or DNA. That is, modified oligonucleotides comprise at least one modified nucleoside (comprising a modified sugar moiety and/or a modified nucleobase) and/or at least one modified intemucleoside linkage.
  • an oligomeric compound comprising a modified oligonucleotide consisting of 17 to 30 linked nucleosides and having a nucleobase sequence comprising at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, or at least 25 consecutive nucleobases of any of the nucleobase sequences of SEQ ID NOs: 19-22 or 63-86, wherein the modified oligonucleotide comprises at least one modification selected from a modified sugar moiety and a modified intemucleoside linkage.
  • the modified oligonucleotide consists of 18-25 linked nucleosides.
  • the modified oligonucleotide consists of 18, 23 or 25 linked nucleosides.
  • Certain embodiments provide an oligomeric compound comprising a modified oligonucleotide comprising the nucleobase sequence of any of SEQ ID Nos: 19-22 or 63-86.
  • Certain embodiments provide an oligomeric compound comprising a modified oligonucleotide consisting of the nucleobase sequence of any of SEQ ID Nos: 19-22 or 63-86.
  • A an adenine nucleobase
  • mC a 5-methylcytosine nucleobase
  • G a guanine nucleobase
  • T a thymine nucleobase
  • n a 2’-NMA sugar moiety
  • s a phosphorothioate intemucleoside linkage
  • o a phosphodiester intemucleoside linkage.
  • Certain embodiments provide an oligomeric compound comprising a modified oligonucleotide according to the following chemical notation: (SEQ ID NO: 45); (SEQ ID NO: 46);
  • A an adenine nucleobase
  • mC a 5-methylcytosine nucleobase
  • G a guanine nucleobase
  • T a thymine nucleobase
  • n a 2’-NMA sugar moiety
  • s a phosphorothioate intemucleoside linkage
  • o a phosphodiester intemucleoside linkage.
  • Modified nucleosides comprise a modified sugar moiety or a modified nucleobase or both a modifed sugar moiety and a modified nucleobase.
  • modified nucleosides comprising the following modified sugar moieties and/or the following modified nucleobases may be incorporated into modified oligonucleotides.
  • modified sugar moieties are non-bicyclic modified sugar moieties comprising a furanosyl ring with one or more substituent groups none of which bridges two atoms of the furanosyl ring to form a bicyclic structure.
  • Such non bridging substituents may be at any position of the furanosyl, including but not limited to substituents at the 2’, 3’, 4’, and/or 5’ positions.
  • 2’-substituent groups suitable for non-bicyclic modified sugar moieties include but are not limited to: 2'-O(CH 2 )2OCH 3 (“MOE” or “O-methoxy ethyl”), and 2’-O-N-methyl acetamide (“NMA”) (see U.S. 6,147,200, Prakash et al., 2003, Org. Lett., 5, 403-6).
  • MOE 2'-O(CH 2 )2OCH 3
  • NMA 2’-O-N-methyl acetamide
  • a “2’-O-N-methyl acetamide nucleoside” or “2’-NMA nucleoside” is shown below:
  • the non-bicyclic modified sugar moiety is a 2 ’-MOE sugar moiety or a 2 ’-NMA sugar moiety.
  • each nucleoside of the modified oligonucleotide comprises a modified sugar moiety.
  • the modified sugar moiety is a 2’-NMA sugar moiety.
  • each nucleoside of the modified oligonucleotide comprises a 2 ’-NMA sugar moiety.
  • modified furanosyl sugar moieties and nucleosides incorporating such modified furanosyl sugar moieties are further defined by isomeric configuration.
  • a 2’-deoxyfuranosyl sugar moiety may be in seven isomeric configurations other than the naturally occurring p-D-deoxyribosyl configuration.
  • modified sugar moieties are described in, e.g., WO 2019/157531, incorporated by reference herein.
  • a 2’-modified sugar moiety has an additional stereocenter at the 2’-position relative to a 2’-deoxyfuranosyl sugar moiety; therefore, such sugar moieties have a total of sixteen possible isomeric configurations.
  • 2’-modified sugar moieties described herein are in the P-D-ribosyl isomeric configuration unless otherwise specified.
  • modified oligonucleotides comprise one or more nucleosides comprising an unmodified nucleobase. In certain embodiments, modified oligonucleotides comprise one or more nucleosides comprising a modified nucleobase. Examples of modified nucleobases include 5-methylcytosine.
  • nucleosides of modified oligonucleotides may be linked together using one or more modified intemucleoside linkages.
  • the two main classes of intemucleoside linking groups are defined by the presence or absence of a phosphoms atom.
  • Modified intemucleoside linkages, compared to naturally occurring phosphate linkages can be used to alter, typically increase, nuclease resistance of the oligonucleotide.
  • intemucleoside linkages having a chiral atom can be prepared as a racemic mixture, or as separate enantiomers. Methods of preparation of phosphorous-containing and non-phosphorous-containing intemucleoside linkages are well known to those skilled in the art.
  • intemucleoside linkages having a chiral center include but are not limited to phosphorothioates.
  • Modified oligonucleotides comprising intemucleoside linkages having a chiral center can be prepared as populations of modified oligonucleotides comprising stereorandom intemucleoside linkages, or as populations of modified oligonucleotides comprising phosphorothioate or other linkages containing chiral centers in particular stereochemical configurations.
  • populations of modified oligonucleotides comprise phosphorothioate intemucleoside linkages wherein all of the phosphorothioate intemucleoside linkages are stereorandom.
  • modified oligonucleotides can be generated using synthetic methods that result in random selection of the stereochemical configuration of each phosphorothioate linkage. Nonetheless, each individual phosphorothioate of each individual oligonucleotide molecule has a defined stereoconfiguration.
  • populations of modified oligonucleotides are enriched for modified oligonucleotides comprising one or more particular phosphorothioate intemucleoside linkages in a particular, independently selected stereochemical configuration.
  • the particular configuration of the particular phosphorothioate linkage is present in at least 65% of the molecules in the population.
  • the particular configuration of the particular phosphorothioate linkage is present in at least 70% of the molecules in the population. In certain embodiments, the particular configuration of the particular phosphorothioate linkage is present in at least 80% of the molecules in the population. In certain embodiments, the particular configuration of the particular phosphorothioate linkage is present in at least 90% of the molecules in the population. In certain embodiments, the particular configuration of the particular phosphorothioate linkage is present in at least 99% of the molecules in the population.
  • modified oligonucleotides can be generated using synthetic methods known in the art, e.g., methods described in Oka et al., JACS 125, 8307 (2003), Wan et al. Nuc. Acid. Res. 42, 13456 (2014), and WO 2017/015555.
  • a population of modified oligonucleotides is enriched for modified oligonucleotides having at least one indicated phosphorothioate in the (.S'p) configuration.
  • a population of modified oligonucleotides is enriched for modified oligonucleotides having at least one phosphorothioate in the (7?p) configuration.
  • modified oligonucleotides comprising (Rp) and/or (.S'p) phosphorothioates comprise one or more of the following formulas, respectively, wherein “B” indicates a nucleobase:
  • chiral intemucleoside linkages of modified oligonucleotides described herein can be stereorandom or in a particular stereochemical configuration.
  • modified oligonucleotides comprise one or more modified nucleosides comprising a modified sugar moiety. In certain embodiments, modified oligonucleotides comprise one or more modified nucleosides comprising a modified nucleobase. In certain embodiments, modified oligonucleotides comprise one or more modified intemucleoside linkage. In such embodiments, the modified, unmodified, and differently modified sugar moieties, nucleobases, and/or intemucleoside linkages of a modified oligonucleotide define a pattern or motif. In certain embodiments, the patterns of sugar moieties, nucleobases, and intemucleoside linkages are each independent of one another.
  • a modified oligonucleotide may be described by its sugar motif, nucleobase motif and/or intemucleoside linkage motif (as used herein, nucleobase motif describes the modifications to the nucleobases independent of the sequence of nucleobases).
  • oligonucleotides comprise one or more type of modified sugar and/or unmodified sugar moiety arranged along the oligonucleotide or region thereof in a defined pattern or sugar motif.
  • sugar motifs include but are not limited to any of the sugar modifications discussed herein.
  • each nucleoside of a modified oligonucleotide, or portion thereof comprises a 2’- substituted sugar moiety, a bicyclic sugar moiety, a sugar surrogate, or a 2’-deoxyribosyl sugar moiety.
  • the 2’-substituted sugar moiety is selected from a 2’-MOE sugar moiety or a 2’-NMA sugar moiety.
  • each nucleoside of a modified oligonucleotide comprises a modified sugar moiety (“fully modified oligonucleotide”).
  • each nucleoside of a fully modified oligonucleotide comprises a 2’-substituted sugar moiety.
  • the 2 ’-substituted sugar moiety is selected from a 2’- MOE sugar moiety or a 2’-NMA sugar moiety.
  • each nucleoside of a fully modified oligonucleotide comprises the same modified sugar moiety (“uniformly modified sugar motif’).
  • the uniformly modified sugar motif is 7 to 20 nucleosides in length.
  • each nucleoside of the uniformly modified sugar motif comprises a 2 ’-substituted sugar moiety.
  • the 2’-substituted sugar moiety is selected from a 2’-MOE sugar moiety or a 2’-NMA sugar moiety.
  • the 2’-substituted sugar moiety is a 2’-NMA sugar moiety.
  • oligonucleotides comprise modified and/or unmodified nucleobases arranged along the oligonucleotide or region thereof in a defined pattern or motif.
  • each nucleobase is modified.
  • none of the nucleobases are modified.
  • each purine or each pyrimidine is modified.
  • each cytosine is modified.
  • some, or all of the cytosine nucleobases in a modified oligonucleotide are 5-methylcytosines.
  • all of the cytosine nucleobases are 5-methylcytosines and all of the other nucleobases of the modified oligonucleotide are unmodified nucleobases.
  • oligonucleotides comprise modified and/or unmodified intemucleoside linkages arranged along the oligonucleotide or region thereof in a defined pattern or motif.
  • each intemucleoside linkage of a modified oligonucleotide is independently selected from a phosphorothioate intemucleoside linkage and phosphodiester intemucleoside linkage.
  • each phosphorothioate intemucleoside linkage is independently selected from a stereorandom phosphorothioate, a (S’p) phosphorothioate, and a (ftp) phosphorothioate.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): soosssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sosossssssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sossossssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sosssossssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sossssossssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sssoosssssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sssssssoosssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sssssssssoossssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sssssssssoossss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sosssssssosssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sossssssssosssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sossssssssssssoss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sssssoosssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssssssssssssooss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage. .
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssssssossssssoss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssssssssosssssoss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssssssssssosssoss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): sssssssssssososs, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssssssssossssosss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssssssssssososss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ossssssssssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): osssssssssssssssssssssssso, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssoosoosoosssssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssoosssssssssssssssssssssssssssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssoosssssssssssssssssssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • modified oligonucleotides have an intemucleoside linkage motif of (5’ to 3’): ssoosoossssssssssssssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage. 4. Certain Lengths
  • oligonucleotides can have any of a variety of ranges of lengths.
  • oligonucleotides consist of X to Y linked nucleosides, where X represents the fewest number of nucleosides in the range and Y represents the largest number nucleosides in the range.
  • X and Y are each independently selected from 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, and 50; provided that X ⁇ Y.
  • oligonucleotides consist of 17 to 18, 17 to 19, 17 to 20, 17 to 21, 17 to 22, 17 to 23, 17 to 24, 17 to 25, 17 to 26, 17 to 27, 17 to 28, 17 to 29, 17 to 30, 18 to 19, 18 to 20, 18 to 21, 18 to 22, 18 to 23, 18 to 24, 18 to 25, 18 to 26, 18 to 27, 18 to 28, 18 to 29, 18 to 30, 19 to 20, 19 to 21, 19 to 22, 19 to 23, 19 to 24, 19 to 25, 19 to 26, 19 to 27, 19 to 28, 19 to 29, 19 to 30, 20 to 21, 20 to 22, 20 to 23, 20 to 24, 20 to 25, 20 to 26, 20 to 27, 20 to 28, 20 to 29, 20 to 30, 21 to 22, 21 to 23, 21 to 24, 21 to 25, 21 to 26, 21 to 27, 21 to 28, 21 to 29, 21 to 30, 22 to 23, 22 to 24, 22 to 25, 22 to 26, 22 to 27, 22 to 28, 22 to 29, 22 to 30, 23 to 24, 23 to 25, 23 to 26, 23 to 27, 23 to 28, 23 to 29, 23 to 30, 24 to 25, 24 to 26, 24 to 27, 24 to 28, 24 to 29, 24 to 30, 25 to 26, 25 to 27, 25 to 28,
  • oligonucleotides consist of 17 linked nucleosides. In certain embodiments, oligonucleotides consist of 18 linked nucleosides. In certain embodiments, oligonucleotides consist of 19 linked nucleosides. In certain embodiments, oligonucleotides consist of 20 linked nucleosides. In certain embodiments, oligonucleotides consist of 23 linked nucleosides. In certain embodiments, oligonucleotides consist of 25 linked nucleosides.
  • Populations of modified oligonucleotides in which all of the modified oligonucleotides of the population have the same molecular formula can be stereorandom populations or chirally enriched populations. All of the chiral centers of all of the modified oligonucleotides are stereorandom in a stereorandom population. In a chirally enriched population, at least one particular chiral center is not stereorandom in the modified oligonucleotides of the population.
  • oligomeric compounds which consist of an oligonucleotide (modified or unmodified) and optionally one or more conjugate groups and/or terminal groups.
  • Conjugate groups consist of one or more conjugate moiety and a conjugate linker which links the conjugate moiety to the oligonucleotide. Conjugate groups may be attached to either or both ends of an oligonucleotide and/or at any internal position. In certain embodiments, conjugate groups are attached to the 2'-position of a nucleoside of a modified oligonucleotide. In certain embodiments, conjugate groups that are attached to either or both ends of an oligonucleotide are terminal groups.
  • conjugate groups or terminal groups are attached at the 3’ and/or 5 ’-end of oligonucleotides. In certain such embodiments, conjugate groups (or terminal groups) are attached at the 3 ’-end of oligonucleotides. In certain embodiments, conjugate groups are attached near the 3 ’-end of oligonucleotides. In certain embodiments, conjugate groups (or terminal groups) are attached at the 5 ’-end of oligonucleotides. In certain embodiments, conjugate groups are attached near the 5 ’-end of oligonucleotides. Examples of terminal groups include but are not limited to conjugate groups, capping groups, phosphate moieties, protecting groups, modified or unmodified nucleosides, and two or more nucleosides that are independently modified or unmodified.
  • oligonucleotides are covalently attached to one or more conjugate groups.
  • conjugate groups modify one or more properties of the attached oligonucleotide, including but not limited to pharmacodynamics, pharmacokinetics, stability, binding, absorption, tissue distribution, cellular distribution, cellular uptake, charge and clearance.
  • conjugation of one or more carbohydrate moieties to a modified oligonucleotide can optimize one or more properties of the modified oligonucleotide.
  • the carbohydrate moiety is attached to a modified subunit of the modified oligonucleotide.
  • the ribose sugar of one or more ribonucleotide subunits of a modified oligonucleotide can be replaced with another moiety, e.g., a non-carbohydrate (preferably cyclic) carrier to which is attached a carbohydrate ligand.
  • a ribonucleotide subunit in which the ribose sugar of the subunit has been so replaced is referred to herein as a ribose replacement modification subunit (RRMS), which is a modified sugar moiety.
  • RRMS ribose replacement modification subunit
  • a cyclic carrier may be a carbocyclic ring system, i.e., one or more ring atoms may be a heteroatom, e.g., nitrogen, oxygen, sulphur.
  • the cyclic carrier may be a monocyclic ring system, or may contain two or more rings, e.g., fused rings.
  • the cyclic carrier may be a fully saturated ring system, or it may contain one or more double bonds.
  • conjugate groups impart a new property on the attached oligonucleotide, e.g. , fluorophores or reporter groups that enable detection of the oligonucleotide.
  • Certain conjugate groups and conjugate moieties have been described previously, for example: cholesterol moiety (Letsinger et al., Proc. Natl. Acad. Sci. USA, 1989, 86, 6553-6556), cholic acid (Manoharan et al., Bioorg. Med. Chem. Lett., 1994, 4, 1053-1060), a thioether, e.g., hexyl-S-tritylthiol (Manoharan et al., Ann. N. Y.
  • Acids Res., 1990, 18, 3777-3783 a polyamine or a polyethylene glycol chain (Manoharan et al., Nucleosides & Nucleotides, 1995, 14, 969-973), or adamantane acetic acid a palmityl moiety (Mishra et al., Biochim. Biophys. Acta, 1995, 1264, 229-237), an octadecylamine or hexylamino-carbonyl-oxycholesterol moiety (Crooke et al., J. Pharmacol. Exp.
  • the conjugate group may comprise a conjugate moiety selected from any of a C22 alkyl, C20 alkyl, C16 alkyl, CIO alkyl, C21 alkyl, C19 alkyl, C18 alkyl, C17 alkyl, C15 alkyl, C14 alkyl, C13 alkyl, C12 alkyl, Cll alkyl, C9 alkyl, C8 alkyl, C7 alkyl, C6 alkyl, C5 alkyl, C22 alkenyl, C20 alkenyl, C16 alkenyl, CIO alkenyl, C21 alkenyl, C19 alkenyl, C18 alkenyl, C17 alkenyl, C15 alkenyl, C14 alkenyl, C13 alkenyl, C12 alkenyl, Cl l alkenyl, C9 alkenyl, C8 alkenyl, C7 alkenyl, C6 alkenyl, or C5 alkyl, C
  • the conjugate group may comprise a conjugate moiety selected from any of a C22 alkyl, C20 alkyl, C16 alkyl, CIO alkyl, C21 alkyl, C19 alkyl, C18 alkyl, C17 alkyl, C15 alkyl, C14 alkyl, C13 alkyl, C12 alkyl, Cll alkyl, C9 alkyl, C8 alkyl, C7 alkyl, C6 alkyl, or C5 alkyl, where the alkyl chain has one or more unsaturated bonds.
  • a conjugate group is a lipid having the following structure:
  • Conjugate moieties include, without limitation, intercalators, reporter molecules, polyamines, polyamides, peptides, carbohydrates (e.g., GalNAc), vitamin moieties, polyethylene glycols, thioethers, polyethers, cholesterols, thiocholesterols, cholic acid moieties, folate, lipids, phospholipids, biotin, phenazine, phenanthridine, anthraquinone, adamantane, acridine, fluoresceins, rhodamines, coumarins, fluorophores, and dyes.
  • intercalators include, without limitation, intercalators, reporter molecules, polyamines, polyamides, peptides, carbohydrates (e.g., GalNAc), vitamin moieties, polyethylene glycols, thioethers, polyethers, cholesterols, thiocholesterols, cholic acid moieties, folate, lipids, phospholipids, bio
  • a conjugate moiety comprises an active drug substance, for example, aspirin, warfarin, phenylbutazone, ibuprofen, suprofen, fen-bufen, ketoprofen, (S)-(+)-pranoprofen, carprofen, dansylsarcosine, 2,3,5- triiodobenzoic acid, fingolimod, flufenamic acid, folinic acid, a benzothiadiazide, chlorothiazide, a diazepine, indo-methicin, a barbiturate, a cephalosporin, a sulfa drug, an antidiabetic, an antibacterial or an antibiotic.
  • an active drug substance for example, aspirin, warfarin, phenylbutazone, ibuprofen, suprofen, fen-bufen, ketoprofen, (S)-(+)-pranoprofen, car
  • Conjugate moieties are attached to oligonucleotides through conjugate linkers.
  • the conjugate linker is a single chemical bond (i.e., the conjugate moiety is attached directly to an oligonucleotide through a single bond).
  • the conjugate linker comprises a chain structure, such as a hydrocarbyl chain, or an oligomer of repeating units such as ethylene glycol, nucleosides, or amino acid units.
  • a conjugate linker comprises pyrrolidine.
  • a conjugate linker comprises one or more groups selected from alkyl, amino, oxo, amide, disulfide, polyethylene glycol, ether, thioether, and hydroxylamino. In certain such embodiments, the conjugate linker comprises groups selected from alkyl, amino, oxo, amide and ether groups. In certain embodiments, the conjugate linker comprises groups selected from alkyl and amide groups. In certain embodiments, the conjugate linker comprises groups selected from alkyl and ether groups. In certain embodiments, the conjugate linker comprises at least one phosphoms moiety. In certain embodiments, the conjugate linker comprises at least one phosphate group. In certain embodiments, the conjugate linker includes at least one neutral linking group.
  • conjugate linkers are bifunctional linking moieties, e.g., those known in the art to be useful for attaching conjugate moieties to compounds, such as the oligonucleotides provided herein.
  • a bifunctional linking moiety comprises at least two functional groups. One of the functional groups is selected to react with a particular site on a compound and the other is selected to react with a conjugate moiety. Examples of functional groups used in a bifunctional linking moiety include but are not limited to electrophiles for reacting with nucleophilic groups and nucleophiles for reacting with electrophilic groups.
  • bifunctional linking moieties comprise one or more groups selected from amino, hydroxyl, carboxylic acid, thiol, alkyl, alkenyl, and alkynyl.
  • conjugate linkers include but are not limited to pyrrolidine, 8-amino-3,6-dioxaoctanoic acid (ADO), succinimidyl 4-(N-maleimidomethyl) cyclohexane- 1 -carboxylate (SMCC) and 6-aminohexanoic acid (AHEX or AHA).
  • ADO 8-amino-3,6-dioxaoctanoic acid
  • SMCC succinimidyl 4-(N-maleimidomethyl) cyclohexane- 1 -carboxylate
  • AHEX or AHA 6-aminohexanoic acid
  • conjugate linkers include but are not limited to substituted or unsubstituted Ci-Cw alkyl, substituted or unsubstituted C2-C10 alkenyl or substituted or unsubstituted C2-C10 alkynyl, wherein a nonlimiting list of preferred substituent groups includes hydroxyl, amino, alkoxy, carboxy, benzyl, phenyl, nitro, thiol, thioalkoxy, halogen, alkyl, aryl, alkenyl and alkynyl.
  • conjugate linkers comprise 1-10 linker-nucleosides. In certain embodiments, conjugate linkers comprise 2-5 linker-nucleosides. In certain embodiments, conjugate linkers comprise exactly 3 linker- nucleosides. In certain embodiments, conjugate linkers comprise the TCA motif. In certain embodiments, such linker- nucleosides are modified nucleosides. In certain embodiments such linker-nucleosides comprise a modified sugar moiety. In certain embodiments, linker-nucleosides are unmodified. In certain embodiments, linker-nucleosides comprise an optionally protected heterocyclic base selected from a purine, substituted purine, pyrimidine or substituted pyrimidine.
  • a cleavable moiety is a nucleoside selected from uracil, thymine, cytosine, 4-N- benzoylcytosine, 5-methylcytosine, 4-N-benzoyl-5-methylcytosine, adenine, 6-N-benzoyladenine, guanine and 2-N- isobutyrylguanine. It is typically desirable for linker-nucleosides to be cleaved from the oligomeric compound after it reaches a target tissue. Accordingly, linker-nucleosides are typically linked to one another and to the remainder of the oligomeric compound through cleavable bonds. In certain embodiments, such cleavable bonds are phosphodiester bonds.
  • linker-nucleosides are not considered to be part of the oligonucleotide. Accordingly, in embodiments in which an oligomeric compound comprises an oligonucleotide consisting of a specified number or range of linked nucleosides and/or a specified percent complementarity to a reference nucleic acid and the oligomeric compound also comprises a conjugate group comprising a conjugate linker comprising linker-nucleosides, those linker-nucleosides are not counted toward the length of the oligonucleotide and are not used in determining the percent complementarity of the oligonucleotide for the reference nucleic acid.
  • an oligomeric compound may comprise (1) a modified oligonucleotide consisting of 8-30 nucleosides and (2) a conjugate group comprising 1-10 linker-nucleosides that are contiguous with the nucleosides of the modified oligonucleotide.
  • the total number of contiguous linked nucleosides in such an oligomeric compound is more than 30.
  • an oligomeric compound may comprise a modified oligonucleotide consisting of 8-30 nucleosides and no conjugate group. The total number of contiguous linked nucleosides in such an oligomeric compound is no more than 30.
  • conjugate linkers comprise no more than 10 linker-nucleosides.
  • conjugate linkers comprise no more than 5 linker- nucleosides. In certain embodiments, conjugate linkers comprise no more than 3 linker-nucleosides. In certain embodiments, conjugate linkers comprise no more than 2 linker-nucleosides. In certain embodiments, conjugate linkers comprise no more than 1 linker-nucleoside.
  • a conjugate group it is desirable for a conjugate group to be cleaved from the oligonucleotide.
  • oligomeric compounds comprising a particular conjugate moiety are better taken up by a particular cell type, but once the oligomeric compound has been taken up, it is desirable that the conjugate group be cleaved to release the unconjugated or parent oligonucleotide.
  • certain conjugate linkers may comprise one or more cleavable moieties.
  • a cleavable moiety is a cleavable bond.
  • a cleavable moiety is a group of atoms comprising at least one cleavable bond.
  • a cleavable moiety comprises a group of atoms having one, two, three, four, or more than four cleavable bonds.
  • a cleavable moiety is selectively cleaved inside a cell or subcellular compartment, such as a lysosome.
  • a cleavable moiety is selectively cleaved by endogenous enzymes, such as nucleases.
  • a cleavable bond is selected from among an amide, an ester, an ether, one or both esters of a phosphodiester, a phosphate ester, a carbamate, or a disulfide. In certain embodiments, a cleavable bond is one or both of the esters of a phosphodiester. In certain embodiments, a cleavable moiety comprises a phosphate or phosphodiester. In certain embodiments, the cleavable moiety is a phosphate linkage between an oligonucleotide and a conjugate moiety or conjugate group.
  • a cleavable moiety comprises or consists of one or more linker-nucleosides.
  • the one or more linker-nucleosides are linked to one another and/or to the remainder of the oligomeric compound through cleavable bonds.
  • such cleavable bonds are unmodified phosphodiester bonds.
  • a cleavable moiety is 2'-deoxynucleoside that is attached to either the 3' or 5'-terminal nucleoside of an oligonucleotide by a phosphate intemucleoside linkage and covalently attached to the remainder of the conjugate linker or conjugate moiety by a phosphate or phosphorothioate linkage.
  • the cleavable moiety is 2'-deoxyadenosine.
  • a conjugate group comprises a cell-targeting moiety.
  • a conjugate group has the general formula: wherein n is from 1 to about 3, m is 0 when n is 1, m is 1 when n is 2 or greater, j is 1 or 0, and k is 1 or 0.
  • n is 1, j is 1 and k is 0. In certain embodiments, n is 1, j is 0 and k is 1. In certain embodiments, n is 1, j is 1 and k is 1. In certain embodiments, n is 2, j is 1 and k is 0. In certain embodiments, n is 2, j is 0 and k is 1. In certain embodiments, n is 2, j is 1 and k is 1. In certain embodiments, n is 3, j is 1 and k is 0. In certain embodiments, n is 3, j is 0 and k is 1. In certain embodiments, n is 3, j is 1 and k is 1. In certain embodiments, n is 3, j is 1 and k is 1. In certain embodiments, n is 3, j is 1 and k is 1.
  • conjugate groups comprise cell-targeting moieties that have at least one tethered ligand.
  • cell-targeting moieties comprise two tethered ligands covalently attached to a branching group.
  • each ligand of a cell-targeting moiety has an affinity for at least one type of receptor on a target cell. In certain embodiments, each ligand has an affinity for at least one type of receptor on the surface of a mammalian liver cell. In certain embodiments, each ligand has an affinity for the hepatic asialoglycoprotein receptor (ASGP-R). In certain embodiments, each ligand is a carbohydrate. In certain embodiments, a conjugate group comprises a cell-targeting conjugate moiety. In certain embodiments, a conjugate group has the general formula: wherein n is from 1 to about 3, m is 0 when n is 1, m is 1 when n is 2 or greater, j is 1 or 0, and k is 1 or 0.
  • n is 1, j is 1 and k is 0. In certain embodiments, n is 1, j is 0 and k is 1. In certain embodiments, n is 1, j is 1 and k is 1. In certain embodiments, n is 2, j is 1 and k is 0. In certain embodiments, n is 2, j is 0 and k is 1. In certain embodiments, n is 2, j is 1 and k is 1. In certain embodiments, n is 3, j is 1 and k is 0. In certain embodiments, n is 3, j is 0 and k is 1. In certain embodiments, n is 3, j is 1 and k is 1. In certain embodiments, n is 3, j is 1 and k is 1. In certain embodiments, n is 3, j is 1 and k is 1.
  • conjugate groups comprise cell-targeting moieties that have at least one tethered ligand.
  • cell-targeting moieties comprise two tethered ligands covalently attached to a branching group.
  • cell-targeting moieties comprise three tethered ligands covalently attached to a branching group.
  • oligomeric compounds comprise one or more terminal groups.
  • oligomeric compounds comprise a stabilized 5’-phosphate.
  • Stabilized 5’-phosphates include, but are not limited to 5 ’-phosphonates, including, but not limited to 5’-vinylphosphonates.
  • terminal groups comprise one or more abasic sugar moieties and/or inverted nucleosides.
  • terminal groups comprise one or more 2’-linked nucleosides or sugar moieties. In certain such embodiments, the 2’-linked group is an abasic sugar moiety.
  • oligomeric compounds and oligomeric duplexes are capable of hybridizing to a target nucleic acid, resulting in at least one antisense activity; such oligomeric compounds and oligomeric duplexes are antisense compounds.
  • antisense compounds have antisense activity when they reduce or inhibit the amount or activity of a target nucleic acid by 25% or more in the standard cell assay. In certain embodiments, antisense compounds selectively affect one or more target nucleic acid.
  • Such antisense compounds comprise a nucleobase sequence that hybridizes to one or more target nucleic acid, resulting in one or more desired antisense activity and does not hybridize to one or more non-target nucleic acid or does not hybridize to one or more non-target nucleic acid in such a way that results in significant undesired antisense activity.
  • hybridization of an antisense compound to a target nucleic acid results in recruitment of a protein that cleaves the target nucleic acid.
  • certain antisense compounds result in RNase H mediated cleavage of the target nucleic acid.
  • RNase H is a cellular endonuclease that cleaves the RNA strand of an RNA:DNA duplex.
  • the DNA in such an RNA:DNA duplex need not be unmodified DNA.
  • described herein are antisense compounds that are sufficiently “DNA-like” to elicit RNase H activity.
  • one or more non-DNA-like nucleoside in the gap of a gapmer is tolerated.
  • an antisense compound or a portion of an antisense compound is loaded into an RNA-induced silencing complex (RISC), ultimately resulting in cleavage of the target nucleic acid.
  • RISC RNA-induced silencing complex
  • certain antisense compounds result in cleavage of the target nucleic acid by Argonaute.
  • Antisense compounds that are loaded into RISC are RNAi compounds. RNAi compounds may be double-stranded (siRNA or dsRNAi) or single-stranded (ssRNA).
  • hybridization of an antisense compound to a target nucleic acid does not result in recruitment of a protein that cleaves that target nucleic acid. In certain embodiments, hybridization of the antisense compound to the target nucleic acid results in alteration of splicing of the target nucleic acid. In certain embodiments, hybridization of an oligomeric compound to a target nucleic acid results in exon inclusion. In certain embodiments, hybridization of an oligomeric compound to a target nucleic acid results in exon exclusion. In certain embodiments, hybridization of an oligomeric compound to a target nucleic acid results in a reduced amount or level of RNA that includes an NIE.
  • hybridization of an oligomeric compound to a target nucleic acid results in an increase in the amount or activity of a target nucleic acid. In certain embodiments, hybridization of an antisense compound to a target nucleic acid results in inhibition of a binding interaction between the target nucleic acid and a protein or other nucleic acid. In certain embodiments, hybridization of an antisense compound to a target nucleic acid results in alteration of translation of the target nucleic acid.
  • Antisense activities may be observed directly or indirectly.
  • observation or detection of an antisense activity involves observation or detection of a change in an amount of a target nucleic acid or protein encoded by such target nucleic acid, a change in the ratio of splice variants of a nucleic acid or protein and/or a phenotypic change in a cell or animal.
  • oligomeric compounds comprise or consist of a modified oligonucleotide comprising a region that is complementary to a target nucleic acid.
  • the target nucleic acid is an endogenous RNA molecule.
  • the target nucleic acid encodes a protein.
  • the target nucleic acid is selected from: a mature mRNA and a pre-mRNA, including intronic, exonic, and untranslated regions.
  • the target RNA is a mature mRNA.
  • the target nucleic acid is a pre-mRNA.
  • the target region is entirely within an intron.
  • the target region spans an intron/exon junction. In certain embodiments, the target region is at least 50% within an intron.
  • oligomeric compounds comprise or consist of a modified oligonucleotide that is complementary to a target nucleic acid encoding SCN1A, or a portion thereof.
  • the SCN1A target nucleic acid has the nucleobase sequence set forth in SEQ ID NO: 1 (the complement of GENBANK Accession No. NC 000002.12 truncated from nucleotides 165982001 to 166152000).
  • the SCN1A target nucleic acid has the nucleobase sequence set forth in SEQ ID NO: 2 (GENBANK Accession No. NM 001165963.2).
  • contacting a cell or subject with an oligomeric compound complementary to SEQ ID NO: 1 or SEQ ID NO: 2 modulates splicing of SCN1A RNA in a cell or a subject. In certain embodiments, contacting a cell or a subject with an oligomeric compound complementary to SEQ ID NO: 1 or SEQ ID NO: 2 increases the amount of SCN1A RNA and/or protein. In certain embodiments, contacting a cell or a subject with an oligomeric compound complementary to SEQ ID NO: 1 or SEQ ID NO: 2 reduces the amount of SCN1A RNA including a NIE.
  • contacting a cell or a subject with an oligomeric compound complementary to SEQ ID NO: 1 or SEQ ID NO: 2 increases the amount of SCN1A RNA excluding a NIE.
  • the NIE is NIE-1.
  • the oligomeric compound comprises or consists of a modified oligonucleotide.
  • contacting a cell in a subject with an oligomeric compound complementary to SEQ ID NO: 1 or SEQ ID NO: 2 ameliorates one or more symptom or hallmark of a disease or disorder associated with SCN1A.
  • the disease or disorder associated with SCN1A is a DEE.
  • the DEE is Dravet Syndrome.
  • the symptom is any of seizures that are prolonged in duration (often lasting longer than 10 minutes), frequent seizures (for example, convulsive, myoclonic, absence, focal, obtundation status, and tonic seizures), sudden unexpected death in epilepsy, status epilepticus, behavioral dysfunctions (for example, aggressiveness, agitation, obsessiveness, preservation, hoarding behavior, or sleep disorders), and developmental delays, movement and balance dysfunctions, orthopedic conditions, motor system and cognitive dysfunctions (for example, ataxia, tremors, dysarthria, pyramidal, and extrapyramidal signs), cognitive impairment, delayed language and speech issues, visual motor integration dysfunctions, visual perception dysfunctions, executive dysfunctions, growth and nutrition issues, sleeping difficulties, chronic infections, sensory integration disorders, and dysautonomia
  • oligomeric compounds comprise or consist of a modified oligonucleotide comprising a portion that is complementary to a target nucleic acid, wherein the target nucleic acid is expressed in a pharmacologically relevant tissue.
  • the pharmacologically relevant tissues are the cells and tissues that comprise the central nervous system (CNS). Such tissues include brain tissues, such as, cerebral cortex, .hippocampus, brainstem, and thalamus.
  • compositions comprising one or more oligomeric compounds.
  • the one or more oligomeric compounds each consists of a modified oligonucleotide.
  • the pharmaceutical composition comprises a pharmaceutically acceptable diluent or carrier.
  • a pharmaceutical composition comprises or consists of a sterile saline solution and one or more oligomeric compound.
  • the sterile saline is pharmaceutical grade saline.
  • a pharmaceutical composition comprises or consists of one or more oligomeric compound and sterile water.
  • the sterile water is pharmaceutical grade water.
  • a pharmaceutical composition comprises or consists of one or more oligomeric compound and phosphate- buffered saline (PBS).
  • the sterile PBS is pharmaceutical grade PBS.
  • a pharmaceutical composition comprises or consists of one or more oligomeric compound and artificial cerebrospinal fluid.
  • the artificial cerebrospinal fluid is pharmaceutical grade artificial cerebrospinal fluid.
  • a pharmaceutical composition comprises a modified oligonucleotide and PBS.
  • a pharmaceutical composition consists of a modified oligonucleotide and PBS.
  • a pharmaceutical composition consists essentially of a modified oligonucleotide and PBS.
  • the PBS is pharmaceutical grade.
  • a pharmaceutical composition comprises a modified oligonucleotide and artificial cerebrospinal fluid. In certain embodiments, a pharmaceutical composition consists of a modified oligonucleotide and artificial cerebrospinal fluid. In certain embodiments, a pharmaceutical composition consists essentially of a modified oligonucleotide and artificial cerebrospinal fluid. In certain embodiments, the artificial cerebrospinal fluid is pharmaceutical grade.
  • compositions comprise one or more oligomeric compound and one or more excipients.
  • excipients are selected from water, salt solutions, alcohol, polyethylene glycols, gelatin, lactose, amylase, magnesium stearate, talc, silicic acid, viscous paraffin, hydroxymethylcellulose and polyvinylpyrrolidone .
  • oligomeric compounds may be admixed with pharmaceutically acceptable active and/or inert substances for the preparation of pharmaceutical compositions or formulations.
  • Compositions and methods for the formulation of pharmaceutical compositions depend on a number of criteria, including, but not limited to, route of administration, extent of disease, or dose to be administered.
  • compositions comprising an oligomeric compound encompass any pharmaceutically acceptable salts of the oligomeric compound, esters of the oligomeric compound, or salts of such esters.
  • pharmaceutical compositions comprising oligomeric compounds comprising one or more oligonucleotide upon administration to an animal, including a human, are capable of providing (directly or indirectly) the biologically active metabolite or residue thereof.
  • the disclosure is also drawn to pharmaceutically acceptable salts of oligomeric compounds, prodrugs, pharmaceutically acceptable salts of such prodrugs, and other bioequivalents.
  • Suitable pharmaceutically acceptable salts include, but are not limited to, sodium, potassium, calcium, and magnesium salts.
  • prodrugs comprise one or more conjugate group attached to a modified oligonucleotide, wherein the conjugate group is cleaved by endogenous nucleases within the body.
  • Lipid moieties have been used in nucleic acid therapies in a variety of methods.
  • the nucleic acid such as an oligomeric compound, is introduced into preformed liposomes or lipoplexes made of mixtures of cationic lipids and neutral lipids.
  • DNA complexes with mono- or poly -cationic lipids are formed without the presence of a neutral lipid.
  • a lipid moiety is selected to increase distribution of a pharmaceutical agent to a particular cell or tissue.
  • a lipid moiety is selected to increase distribution of a pharmaceutical agent to fat tissue.
  • a lipid moiety is selected to increase distribution of a pharmaceutical agent to muscle tissue.
  • compositions comprise a delivery system.
  • delivery systems include, but are not limited to, liposomes and emulsions. Certain delivery systems are useful for preparing certain pharmaceutical compositions including those comprising hydrophobic compounds.
  • certain organic solvents such as dimethylsulfoxide are used.
  • pharmaceutical compositions comprise one or more tissue-specific delivery molecules designed to deliver the one or more pharmaceutical agents comprising an oligomeric compound provided herein to specific tissues or cell types.
  • pharmaceutical compositions include liposomes coated with a tissue-specific antibody.
  • compositions comprise a co-solvent system.
  • co-solvent systems comprise, for example, benzyl alcohol, a nonpolar surfactant, a water-miscible organic polymer, and an aqueous phase.
  • co-solvent systems are used for hydrophobic compounds.
  • a non-limiting example of such a co-solvent system is the VPD co-solvent system, which is a solution of absolute ethanol comprising 3% w/v benzyl alcohol, 8% w/v of the nonpolar surfactant Polysorbate 80TM and 65% w/v polyethylene glycol 300.
  • the proportions of such co-solvent systems may be varied considerably without significantly altering their solubility and toxicity characteristics.
  • co-solvent components may be varied: for example, other surfactants may be used instead of Polysorbate 80TM; the fraction size of polyethylene glycol may be varied; other biocompatible polymers may replace polyethylene glycol, e.g., polyvinyl pyrrolidone; and other sugars or polysaccharides may substitute for dextrose.
  • compositions are prepared for oral administration.
  • pharmaceutical compositions are prepared for buccal administration.
  • a pharmaceutical composition is prepared for administration by injection (e.g., intravenous, subcutaneous, intramuscular, intrathecal (IT), intracerebroventricular (ICV), etc.).
  • a pharmaceutical composition comprises a carrier and is formulated in aqueous solution, such as water or physiologically compatible buffers such as Hanks's solution, Ringer's solution, or physiological saline buffer.
  • other ingredients are included (e.g., ingredients that aid in solubility or serve as preservatives).
  • injectable suspensions are prepared using appropriate liquid carriers, suspending agents and the like.
  • compositions for injection are presented in unit dosage form, e.g., in ampoules or in multi-dose containers.
  • Certain pharmaceutical compositions for injection are suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
  • Certain solvents suitable for use in pharmaceutical compositions for injection include, but are not limited to, lipophilic solvents and fatty oils, such as sesame oil, synthetic fatty acid esters, such as ethyl oleate or triglycerides, and liposomes.
  • certain compounds disclosed herein act as acids. Although such compounds may be drawn or described in protonated (free acid) form or ionized and in association with a cation (salt) form, aqueous solutions of such compounds exist in equilibrium among such forms. For example, a phosphate linkage of an oligonucleotide in aqueous solution exists in equilibrium among free acid, anion and salt forms. Unless otherwise indicated, compounds described herein are intended to include all such forms. Moreover, certain oligonucleotides have several such linkages, each of which is in equilibrium. Thus, oligonucleotides in solution exist in an ensemble of forms at multiple positions all at equilibrium. The term “oligonucleotide” is intended to include all such forms.
  • a structure depicting the free acid of a compound followed by the term “or a pharmaceutically acceptable salt thereof’ expressly includes all such forms that may be fully or partially protonated/de-protonated/in association with one or more cations selected from sodium, potassium, calcium, and magnesium.
  • modified oligonucleotides or oligomeric compounds are in aqueous solution with sodium. In certain embodiments, modified oligonucleotides or oligomeric compounds are in aqueous solution with potassium. In certain embodiments, modified oligonucleotides or oligomeric compounds are in PBS. In certain embodiments, modified oligonucleotides or oligomeric compounds are in water. In certain such embodiments, the pH of the solution is adjusted with NaOH and/or HC1 to achieve a desired pH.
  • a dose may be in the form of a dosage unit.
  • a dose (or dosage unit) of a modified oligonucleotide or an oligomeric compound in milligrams indicates the mass of the free acid form of the modified oligonucleotide or oligomeric compound.
  • the free acid is in equilibrium with anionic and salt forms.
  • the modified oligonucleotide or oligomeric compound exists as a solvent-free, sodium-acetate free, anhydrous, free acid.
  • a modified oligonucleotide or an oligomeric compound may be partially or fully de-protonated and in association with Na+ ions.
  • the mass of the protons are nevertheless counted toward the weight of the dose, and the mass of the Na+ ions are not counted toward the weight of the dose.
  • a dose, or dosage unit, of 10 mg of Compound No. 1464713 equals the number of fully protonated molecules that weighs 10 mg. This would be equivalent to 10.51 mg of solvent-free, sodium acetate-free, anhydrous sodiated Compound No.
  • an oligomeric compound comprises a conjugate group
  • the mass of the conjugate group is included in calculating the dose of such oligomeric compound. If the conjugate group also has an acid, the conjugate group is likewise assumed to be fully protonated for the purpose of calculating dose.
  • an oligomeric compound may be partially or fully de-protonated and in association with sodium, potassium, calcium, and/or magnesium.
  • the mass of the protons is nevertheless counted toward the weight of the dose, and the mass of the sodium, potassium, calcium, and magnesium ions is not counted toward the weight of the dose.
  • an oligomeric compound comprises a conjugate group
  • the mass of the conjugate group is included in calculating the dose of such oligomeric compound. If the conjugate group also has an acid, the conjugate group is likewise assumed to be fully protonated for the purpose of calculating dose.
  • an oligonucleotide comprising a nucleoside comprising a 2’-OH sugar moiety and a thymine base could be described as a DNA having a modified sugar (2 ’-OH in place of one 2’-H of DNA) or as an RNA having a modified base (thymine (methylated uracil) in place of a uracil of RNA).
  • nucleic acid sequences provided herein, including, but not limited to those in the sequence listing are intended to encompass nucleic acids containing any combination of natural or modified RNA and/or DNA, unless otherwise stated, including, but not limited to such nucleic acids having modified nucleobases.
  • an oligomeric compound having the nucleobase sequence “ATCGATCG” encompasses any oligomeric compounds having such nucleobase sequence, whether modified or unmodified, including, but not limited to, such compounds comprising RNA bases, such as those having sequence “AUCGAUCG” and those having some DNA bases and some RNA bases such as “AUCGATCG” and oligomeric compounds having other modified nucleobases, such as “AT m CGAUCG,” wherein m C indicates a cytosine base comprising a methyl group at the 5-position.
  • Certain compounds described herein e.g., modified oligonucleotides have one or more asymmetric center and thus give rise to enantiomers, diastereomers, and other stereoisomeric configurations that may be defined, in terms of absolute stereochemistry, as (R) or (S), as a or 0 such as for sugar anomers, or as (D) or (L), such as for amino acids, etc.
  • Compounds provided herein that are drawn or described as having certain stereoisomeric configurations include only the indicated compounds.
  • Compounds provided herein that are drawn or described with undefined stereochemistry include all such possible isomers, including their stereorandom and optically pure forms, unless specified otherwise.
  • tautomeric forms of the compounds herein are also included unless otherwise indicated. Unless otherwise indicated, compounds described herein are intended to include corresponding salt forms.
  • the compounds described herein include variations in which one or more atoms are replaced with a nonradioactive isotope or radioactive isotope of the indicated element.
  • compounds herein that comprise hydrogen atoms encompass all possible deuterium substitutions for each of the 1 H hydrogen atoms.
  • Isotopic substitutions encompassed by the compounds herein include but are not limited to: 2 H or 3 H in place of 'H. 13 C or 14 C in place of 12 C, 15 N in place of 14 N, 17 O or 18 O in place of 16 O, and 33 S, 34 S, 35 S, or 36 S in place of 32 S.
  • non-radioactive isotopic substitutions may impart new properties on the oligomeric compound that are beneficial for use as a therapeutic or research tool.
  • radioactive isotopic substitutions may make the compound suitable for research or diagnostic purposes such as imaging.
  • Modified oligonucleotides complementary to a human SCN1A nucleic acid were designed and synthesized as indicated in the table below.
  • Each modified oligonucleotide listed in the tables below is 100% complementary to the human SCN1A genomic sequence, designated herein as SEQ ID NO: 1 (the complement of GENBANK Accession No. NC 000002.12 truncated from nucleotides 165982001 to 166152000), and to the mouse SCN1A genomic sequence, designated herein as SEQ ID NO: 3 (the complement of GENBANK Accession No. NC 000068.7 truncated from nucleotides 66268001 to 66444000).
  • Start site indicates the 5’-most nucleoside to which the modified oligonucleotide is complementary in the target nucleic acid sequence.
  • “Stop site” indicates the 3 ’-most nucleoside to which the modified oligonucleotide is complementary in the target nucleic acid sequence.
  • the modified oligonucleotides in the table below are 18 nucleosides in length.
  • the sugar motif for the modified oligonucleotides in the table below are (from 5’ to 3’): nrmrmrmrmrmrmnnnnn, wherein each “n” represents a 2’-NMA sugar moiety.
  • the intemucleoside linkage motif for each modified oligonucleotide is provided in the Intemucleoside Linkage Motif (5’ to 3’)” column in the table below, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • Each cytosine residue is a 5-methylcytosine.
  • the modified oligonucleotide in the table below is 19 nucleosides in length.
  • the sugar motif for the modified oligonucleotide is (from 5’ to 3’): nnnnnnnnnnnnnnnnnnn, wherein each “n” represents a 2’-NMA sugar moiety.
  • the intemucleoside linkage motif for the modified oligonucleotide is (from 5’to 3’): osssssssssssssssssss, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • Each cytosine residue is a 5-methylcytosine.
  • the modified oligonucleotide in the table below is 20 nucleosides in length.
  • the sugar motif for the modified oligonucleotide is (from 5’ to 3’): nnnnnnnnnnnnnnnnnnnnnnnnn, wherein each “n” represents a 2’-NMA sugar moiety.
  • the intemucleoside linkage motif for the modified oligonucleotide is (from 5’to 3’): ossssssssssssssssssssso, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage Each cytosine residue is a 5-methylcytosine.
  • Example 2 Effect of modified oligonucleotides targeting SCN1A in wildtype mice
  • Wildtype C57BL/6 female mice were divided into groups of 3 mice each. Each mouse received a single ICV bolus of 50 pg of modified oligonucleotide. A group of 4 mice received PBS as a negative control.
  • Compound No. 1429226 is a modified oligonucleotide having a nucleobase sequence of (from 5’ to 3’) AGTTGGAGCAAGATTATC (SEQ ID NO: 13), wherein each nucleoside comprises a 2’-NMA sugar moiety, each intemucleoside linkage is a phosphorothioate intemucleoside linkage, and each cytosine is a 5-methylcytosine.
  • Comparator compound 1367010 has the nucleobase sequence, sugar motif, and intemucleoside linkage motif of Compound Ex 20X+1, previously described in WO 2019/040923 (incorporated herein by reference).
  • Comparator compound 1367010 has a nucleobase sequence of (from 5’ to 3’) AGTTGGAGCAAGATTATC (SEQ ID NO: 13), wherein each nucleoside comprises a 2 ’-MOE sugar moiety, and each intemucleoside linkage is a phosphorothioate intemucleoside linkage.
  • Each cytosine in Comparator compound 1367010 is a 5-methylcytosine.
  • SEQ ID NO: 13 is 100% complementary to SEQ ID NO: 1, from Start Site 144708 to Stop Site 144725, and is 100% complementary to SEQ ID NO: 3 from Start Site 150106 to Stop Site 150123.
  • Start site indicates the 5’-most nucleoside to which the modified oligonucleotide is complementary in the target nucleic acid sequence.
  • “Stop site” indicates the 3 ’-most nucleoside to which the modified oligonucleotide is complementary in the target nucleic acid sequence.
  • mice Two weeks post treatment, mice were sacrificed and RNA was extracted from cortical brain tissue for real-time qPCR analysis of SCN1 A RNA using mouse primer probe set RTS48951 (forward sequence CCCTAAGAGCCTTATCACGATTT, designated herein as SEQ ID NO: 4; reverse sequence GGCAAACCAGAAGCACATTC, designated herein as SEQ ID NO: 5; probe sequence AGGGTGGTTGTGAATGCCCTGTTA, designated herein as SEQ ID NO: 6) to measure the amount of SCN1A RNA that excludes the mouse form of NIE-1 (NIE-T), and mouse primer probe set RTS48949 (forward sequence AGCCCTTTATTATGGGTGGTT, designated herein as SEQ ID NO: 7; reverse sequence CCAGAATATAAGGCAAACCAGAAG, designated herein as SEQ ID NO: 8; probe sequence TGGATGGAATTGCTCCTAACAGGGC, designated herein as SEQ ID NO: 9) to measure the amount of SCN1A transcript that includes the mouse form of
  • SCN1A RNA is presented as the percent of SCN1A RNA relative to the average of the amount in PBS treated animals (%control), normalized to mouse GAPDH.
  • Mouse GAPDH was amplified using primer probe set mGapdh_LTS00102 (forward sequence GGCAAATTCAACGGCACAGT, designated herein as SEQ ID NO: 10; reverse sequence GGGTCTCGCTCCTGGAAGAT, designated herein as SEQ ID NO: 11; probe sequence AAGGCCGAGAATGGGAAGCTTGTCATC, designated herein as SEQ ID NO: 12).
  • Example 3 Tolerability of modified oligonucleotides complementary to SCN1A in wild-type mice
  • Modified oligonucleotides described above were tested in wild-type mice to assess the tolerability of the oligonucleotides.
  • Wild-type female C57/B16 mice each received a single ICV dose of 700 pg of modified oligonucleotide.
  • Each treatment group consisted of 3-4 mice.
  • a group of 4 mice received PBS as a negative control.
  • mice were evaluated according to seven different criteria. The criteria are: (1) the mouse was bright, alert, and responsive; (2) the mouse was standing or hunched without stimuli; (3) the mouse showed any movement without stimuli; (4) the mouse demonstrated forward movement after it was lifted; (5) the mouse demonstrated any movement after it was lifted; (6) the mouse responded to tail pinching; (7) regular breathing.
  • a mouse was given a subscore of 0 if it met the criteria and 1 if it did not (the functional observational battery score or FOB). After all 7 criteria were evaluated, the scores were summed for each mouse and averaged within each treatment group.
  • Example 4 Tolerability of modified oligonucleotides complementary to human SCN1A in rats, 3-hour study
  • oligonucleotides described above were tested in rats to assess the tolerability of the oligonucleotides.
  • Sprague Dawley rats each received a single intrathecal (IT) dose of 3 mg of oligonucleotide listed in the table below.
  • Each treatment group consisted of 3-4 rats.
  • a group of 4 rats received PBS as a negative control.
  • movement in 7 different parts of the body were evaluated for each rat.
  • the 7 body parts are: (1) the rat’s tail; (2) the rat’s posterior posture; (3) the rat’s hind limbs; (4) the rat’s hind paws; (5) the rat’s forepaws; (6) the rat’s anterior posture; (7) the rat’s head.
  • each rat was given a sub-score of 0 if the body part was moving or 1 if the body part was paralyzed (the functional observational battery score or FOB). After each of the 7 body parts were evaluated, the sub-scores were summed for each rat and then averaged for each group.
  • Modified oligonucleotides complementary to a SCN1A nucleic acid were designed and synthesized as indicated in the tables below.
  • the modified oligonucleotides listed in the table below are 100% complementary to the human SCN1A genomic sequence, designated herein as SEQ ID NO: 1 (described herein above), and to the mouse SCN1A genomic sequence, designated herein as SEQ ID NO: 3 (described herein above).
  • Start site indicates the 5’-most nucleoside to which the modified oligonucleotide is complementary in the target nucleic acid sequence.
  • Stop site indicates the 3 most nucleoside to which the modified oligonucleotide is complementary in the target nucleic acid sequence.
  • N. A.” indicates that the modified oligonucleotide is not 100% complementary to the target nucleic acid sequence.
  • the modified oligonucleotides in the table below are 18 nucleosides in length.
  • the sugar motif for the modified oligonucleotides in the table below are (from 5’ to 3’): nnnnnnnnnnnnnnnn, wherein each “n” represents a 2’-NMA sugar moiety.
  • the intemucleoside linkage motif for the modified oligonucleotides is (from 5 ’ to 3 ’): ssssssssssssssssssssss; wherein each “s” represents a phosphorothioate intemucleoside linkage.
  • Each cytosine residue is a 5-methylcytosine.
  • the modified oligonucleotides in the table below are 25 nucleosides in length.
  • the sugar motif for the modified oligonucleotides in the table below are (from 5’ to 3’): nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn, wherein each “n” represents a 2’- NMA sugar moiety.
  • the intemucleoside linkage motif for the modified oligonucleotides is (from 5’ to 3’): ssoosoosoosssssssssssss; wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • Each cytosine residue is a 5-methylcytosine.
  • the modified oligonucleotides in the table below are 25 nucleosides in length.
  • the sugar motif for the modified oligonucleotides in the table below are (from 5’ to 3’): nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn, wherein each “n” represents a 2’- NMA sugar moiety.
  • the intemucleoside linkage motif for the modified oligonucleotides is (from 5’ to 3’): ssoosoosssssssssssssssssss; wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • Each cytosine residue is a 5-methylcytosine.
  • the modified oligonucleotides in the table below are 25 nucleosides in length.
  • the sugar motif for the modified oligonucleotides in the table below are (from 5’ to 3’): nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn, wherein each “n” represents a 2’- NMA sugar moiety.
  • the intemucleoside linkage motif for the modified oligonucleotides is (from 5’ to 3’): ssoosssssssssssssssssssssssssssss; wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • Each cytosine residue is a 5-methylcytosine.
  • the modified oligonucleotides in the table below are 25 nucleosides in length.
  • the sugar motif for the modified oligonucleotides in the table below are (from 5’ to 3’): nnnnnnnnnnnnnnnnnnnnnnnnnnnnnn, wherein each “n” represents a 2’- NMA sugar moiety.
  • the intemucleoside linkage motif for the modified oligonucleotides is (from 5’ to 3’): ssssssssssssssssssssssssssssssssss; wherein each “s” represents a phosphorothioate intemucleoside linkage.
  • Each cytosine residue is a 5-methylcytosine.
  • the modified oligonucleotides in the table below are 23 nucleosides in length.
  • the sugar motif for the modified oligonucleotides in the table below are (from 5’ to 3’): nnnnnnnnnnnnnnnnnnnnnnnnnnnnnn, wherein each “n” represents a 2’- NMA sugar moiety.
  • the intemucleoside linkage motifs for the modified oligonucleotides are presented in the column labeled “Intemucleoside Linkages (5’ to 3’)” in the table below, wherein each “s” represents a phosphorothioate intemucleoside linkage and each “o” represents a phosphodiester intemucleoside linkage.
  • Each cytosine residue is a 5- methylcytosine.
  • “Start site” indicates the 5 ’-most nucleoside to which the modified oligonucleotide is complementary in the target nucleic acid sequence. “Stop site” indicates the 3 ’-most nucleoside to which the modified oligonucleotide is complementary in the target nucleic acid sequence.
  • the modified oligonucleotides listed in the table below are 100% complementary to the mouse SCN1A sequence designated herein as SEQ ID NO: 3 (described herein above); the sequences are complementary to the human SCN1A sequence of SEQ ID NO: 1 (described herein above) with a single mismatch located at the position indicated in the column labeled “Position of mismatch on Compound (5’ to 3’)”.
  • the non-complementary nucleobases are marked in the Nucleobase Sequence column in underlined, bold, italicized font. Additionally, the modified oligonucleotides listed in the table below are 100% complementary to the mouse SCN1A genomic sequence, designated herein as SEQ ID NO: 3 (described herein above).
  • the modified oligonucleotides in the table below are 18 nucleosides in length.
  • the sugar motif for the modified oligonucleotides in the table below are (from 5’ to 3’): nnnnnnnnnnnnnnnn, wherein each “n” represents a 2’-NMA sugar moiety.
  • the intemucleoside linkage motif for the modified oligonucleotides is (from 5 ’ to 3 ’): ssssssssssssssssssssss; wherein each “s” represents a phosphorothioate intemucleoside linkage.
  • Each cytosine residue is a 5-methylcytosine.
  • Example 6 Effect of modified oligonucleotides targeting SCN1A in wildtype mice
  • Wildtype C57BL/6 mice were divided into groups of 3 mice each. Each mouse received a single ICV bolus of 50 pg of modified oligonucleotide. A group of 4 mice received PBS as a negative control.
  • mice Two weeks post treatment, mice were sacrificed, and RNA was extracted from cortical brain tissue for realtime qPCR analysis of SCN 1 A RNA using mouse primer probe set RTS48951 (described herein above) to measure the amount of SCN1A RNA that excludes the mouse form of NIE-1 (NIE- 1'). and mouse primer probe set RTS48949 (described herein above) to measure the amount of SCN1A transcript that includes the mouse form of NIE-1 (NIE-1 + ).
  • SCN 1 A RNA is presented as the percent of SCN1 A RNA relative to the average of the amount in PBS treated animals (%control), normalized to mouse GAPDH.
  • Mouse GAPDH was amplified using primer probe set mGapdh_LTS00102 (described herein above). Values marked with a “f ” result from oligonucleotides that are complementary to the amplicon region of the primer probe set. Additional assays may be used to measure the potency and efficacy of the modified oligonucleotides complementary to the amplicon region.
  • Wildtype C57BL/6 female mice were divided into groups of 4 mice each. Each mouse received a single ICV bolus of modified oligonucleotide at various doses defined in the tables below. A group of 4 mice received PBS as a negative control. Two weeks post treatment, mice were sacrificed, and RNA was extracted from cortical brain tissue for realtime qPCR analysis of SCN 1 A RNA using mouse primer probe set RTS48951 ( described herein above) to measure the amount of SCN1A RNA that excludes the mouse form of NIE-1 (NIE-T), and mouse primer probe set RTS48949 (described herein above) to measure the amount of SCN1A transcript that includes the mouse form of NIE-1 (NIE-1 + ).
  • SCN 1 A RNA is presented as the percent of SCN1 A RNA relative to the average of the amount in PBS treated animals (%control), normalized to mouse GAPDH.
  • Mouse GAPDH was amplified using primer probe set mGapdh_LTS00102 (described herein above).
  • ED50s were calculated in using GraphPad Prism.
  • Comparator compound 1367010 is described herein above.
  • Modified oligonucleotides described above were tested in wild-type mice to assess the tolerability of the oligonucleotides.
  • a mouse was given a subscore of 0 if it met the criteria and 1 if it did not (the functional observational battery score or FOB). After all 7 criteria were evaluated, the scores were summed for each mouse and averaged within each treatment group.
  • Example 9 Tolerability of modified oligonucleotides complementary to human SCN1A in rats, 3-hour study Modified oligonucleotides described above were tested in rats to assess the tolerability of the oligonucleotides.
  • Sprague Dawley rats each received a single intrathecal (IT) dose of 3 mg of oligonucleotide as indicated in the tables below.
  • Each treatment group consisted of 4 rats.
  • a group of 4 rats received PBS as a negative control.
  • movement in 7 different parts of the body were evaluated for each rat.
  • the 7 body parts are (1) the rat’s tail; (2) the rat’s posterior posture; (3) the rat’s hind limbs; (4) the rat’s hind paws; (5) the rat’s forepaws; (6) the rat’s anterior posture; (7) the rat’s head.
  • each rat was given a sub-score of 0 if the body part was moving or 1 if the body part was paralyzed (the functional observational battery score or FOB).
  • FOB functional observational battery score
  • the sub-scores were summed for each rat and then averaged for each group. For example, if a rat’s tail, head, and all other evaluated body parts were moving 3 hours after the 3 mg IT dose, it would get a summed score of 0. If another rat was not moving its tail 3 hours after the 3 mg IT dose but all other evaluated body parts were moving, it would receive a score of 1. Results are presented as the average score for each treatment group. Table 28

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US12338437B2 (en) 2020-05-11 2025-06-24 Stoke Therapeutics, Inc. OPA1 antisense oligomers for treatment of conditions and diseases
WO2026052819A1 (en) * 2024-09-06 2026-03-12 Ospedale San Raffaele S.R.L. Gene therapy
US12577561B2 (en) 2019-02-27 2026-03-17 Stoke Therapeutics, Inc. Antisense oligomers for treatment of conditions and diseases

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US20220372489A1 (en) * 2019-06-21 2022-11-24 Quralis Corporation Ppm1a inhibitors and methods of using same
EP4013767A4 (en) * 2019-08-15 2023-10-25 Ionis Pharmaceuticals, Inc. Linkage modified oligomeric compounds and uses thereof
BR112022010882A2 (pt) * 2019-12-06 2022-10-04 Stoke Therapeutics Inc Oligômeros antissenso para tratamento de condições e doenças
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US12577561B2 (en) 2019-02-27 2026-03-17 Stoke Therapeutics, Inc. Antisense oligomers for treatment of conditions and diseases
US12338437B2 (en) 2020-05-11 2025-06-24 Stoke Therapeutics, Inc. OPA1 antisense oligomers for treatment of conditions and diseases
WO2025024568A1 (en) * 2023-07-24 2025-01-30 Stoke Therapeutics, Inc. Methods for treating conditions and diseases
WO2026052819A1 (en) * 2024-09-06 2026-03-12 Ospedale San Raffaele S.R.L. Gene therapy

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