WO2022047105A2 - Nucleic acids for inhibiting tert expression - Google Patents

Nucleic acids for inhibiting tert expression Download PDF

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WO2022047105A2
WO2022047105A2 PCT/US2021/047864 US2021047864W WO2022047105A2 WO 2022047105 A2 WO2022047105 A2 WO 2022047105A2 US 2021047864 W US2021047864 W US 2021047864W WO 2022047105 A2 WO2022047105 A2 WO 2022047105A2
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aso
nucleic acid
oligonucleotide
sequence
nucleotides
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WO2022047105A3 (en
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Robert Joseph Allen BELL
Jasmin GEGNER
Jochen DECKERT
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Telo Therapeutics, Inc.
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    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
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    • C12N2310/00—Structure or type of the nucleic acid
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    • C12N2310/11—Antisense
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    • C12N2310/00—Structure or type of the nucleic acid
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    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
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    • C12N2310/315—Phosphorothioates
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    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/32—Chemical structure of the sugar
    • C12N2310/323—Chemical structure of the sugar modified ring structure
    • C12N2310/3231—Chemical structure of the sugar modified ring structure having an additional ring, e.g. LNA, ENA
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    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/30—Chemical structure
    • C12N2310/34—Spatial arrangement of the modifications
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    • C12N2320/30—Special therapeutic applications
    • C12N2320/33—Alteration of splicing

Definitions

  • the disclosure relates to compositions and methods for treating cancer by downregulating telomerase.
  • the disclosure provides nucleic acids that inhibit GABPB1 expression (e.g., partial GABPB1 expression or total GABPB1 expression).
  • the nucleic acids are oligonucleotides or siRNAs that target exon 9 of GABPB1L mRNA and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations.
  • the nucleic acids are oligonucleotides or siRNAs that target the intron between exon 8 and exon 9 of GABPB1L mRNA and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations.
  • the nucleic acids are oligonucleotides or siRNAs that target total GABPB1 by targeting both GABPB1S and GABPB1L isoforms of the GABPB1 gene, and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations.
  • BACKGROUND [0004] Telomerase expression is a hallmark of tumorigenesis.
  • telomere Due to its fundamental nature in driving tumorigenesis, many attempts have been made to inhibit telomerase as a cancer therapeutic strategy, but thus far none have become a standard of care.
  • GRN163L By hybridizing and inhibiting the RNA template of telomerase, GRN163L reduced tumor growth in preclinical models of breast cancer, glioblastoma (GBM), pancreatic, and liver cancer. This preclinical success has not translated to the clinic however, as trials in breast, lung, and pediatric CNS cancers were discontinued. In each case, a high frequency of grade III/IV hematopoietic toxicities were observed.
  • the present disclosure provides compositions and methods for treating cancer by downregulating telomerase.
  • the disclosure provides nucleic acids that inhibit GABPB1 expression (e.g., partial GABPB1 expression or total GABPB1 expression).
  • the nucleic acids are oligonucleotides or siRNAs that target exon 9 of GABPB1L mRNA and reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations. In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target the intron between exon 8 and exon 9 of GABPB1L mRNA and reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations.
  • the nucleic acids are oligonucleotides or siRNAs that target total GABPB1 by targeting both GABPB1S and GABPB1L isoforms of the GABPB1 gene, and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations.
  • the present disclosure provides an oligonucleotide capable of hybridizing to a target sequence.
  • the target sequence may comprise exon 9 or the intron between exon 8 and 9 of a GABPB1 gene (DNA) or gene product (RNA) and the target sequence may be, but not limited to, SEQ ID NOs: 308-346, and 12424-24399.
  • the oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 50% of the oligonucleotide sequence.
  • the oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 75% of the oligonucleotide sequence.
  • the oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 85% of the oligonucleotide sequence.
  • the oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 90% of the oligonucleotide sequence.
  • the oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 99% of the oligonucleotide sequence.
  • the oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over 100% of the oligonucleotide sequence.
  • the oligonucleotide may include one or more modified nucleobases, sugars and/or linkers such as, but not limited to, LNAs, phosphorothioate (Ps) linkages, phosphodiester (Po) linkages, 5- methylcytosine modified nucleotides, and 2’MOE modified nucleotides.
  • the present disclosure provides a double-stranded small inhibitory RNA (siRNA) comprising the sequence of any one of SEQ ID NO: 386-424.
  • siRNA a double-stranded small inhibitory RNA
  • the present disclosure provides a method of reducing the activity of GA binding protein transcription factor subunit beta 1 (GABPB1) in a cell, comprising exposing the cell to any of the oligonucleotides, siRNA, ASOs, and pharmaceutical compositions thereof which are disclosed herein.
  • the activity of GABPB1L isoform, GABPB1S isoform or total GABPB1 may be reduced.
  • the present disclosure provides a method of reducing the expression of telomerase reverse transcriptase (TERT) in a cell having a mutant TERT promoter with one or more somatic mutations, comprising exposing the cell to any of the oligonucleotides, siRNA, ASOs, and pharmaceutical compositions thereof which are disclosed herein.
  • the present disclosure provides a pharmaceutical composition, comprising any of the oligonucleotides, siRNA or ASOs and a pharmaceutically-acceptable carrier.
  • the present disclosure provides a medicament, comprising any of the oligonucleotides, siRNA or ASOs and a pharmaceutically-acceptable carrier.
  • the present disclosure provides a method of treating cancer in a subject comprising administering the pharmaceutical compositions described herein to the subject.
  • the present disclosure provides a use of any of the siRNA disclosed herein for the manufacture of a pharmaceutical composition for treating cancer.
  • the present disclosure provides any of the siRNA disclosed herein for use in treating cancer.
  • the present disclosure provides an antisense oligonucleotide (ASO) which may include any of SEQ ID NO: 19-111, 430-474, 12190-12339, and 12361- 12423.
  • ASO antisense oligonucleotide
  • the ASO may comprise SEQ ID NO: 12335, 12338, 12363, 12372, 12376, 12379, 12382, 12383, 12384, 12389, 12393, 12394, 12395, 12413, 12421, and 12422.
  • the present disclosure provides a method of reducing the activity of GA binding protein transcription factor beta 1 (GABPB1) in a cell, comprising exposing the cell to any of the ASOs described herein.
  • the activity of GABPB1L isoform, GABPB1S isoform or total GABPB1 may be reduced.
  • the present disclosure provides a method of reducing the expression of telomerase reverse transcriptase (TERT) in a cell having a mutant TERT promoter with one or more somatic mutations, comprising exposing the cell to any of the ASOs described herein.
  • the present disclosure provides a pharmaceutical composition, comprising any of the ASOs described herein and a pharmaceutically- acceptable carrier.
  • the present disclosure provides a medicament, comprising any of the ASOs described herein and a pharmaceutically-acceptable carrier.
  • the present disclosure provides a method of treating a patient with cancer, comprising administering any of the pharmaceutical compositions described herein to the patient.
  • the present disclosure provides a use of any of the ASOs described herein for the manufacture of a pharmaceutical composition for treating cancer.
  • the present disclosure provides the ASOs described herein for use in treating cancer.
  • DESCRIPTION OF THE DRAWINGS [0022]
  • Figure 1 shows the exon-intron structure of GABPB1L and GABPB1S (not to scale). The inset shows exon 9 of GABPB1L as well as the flanking intron and UTR sequences. Putative splice motifs identified by computational analysis are represented by (*). The tiling window is defined by the vertical dotted lines, and candidate oligonucleotide sequences are represented by the horizontal black lines.
  • FIG. 2 provides a set of charts showing the expression fold change, nano luciferase and fluorescence (590 nm) for the samples.
  • LN229 GBM cells (TERTp mutant) and LN229-P2A-NLuc cells were transfected with 50nM of candidate siRNA.
  • 72 hours post drug treatment cell lysates were harvested and TERT mRNA was measured via RT-qPCR (ThermoFisher, Waltham, MA, Cat. No. A25600).
  • FIG. 3 shows GABPB1L Expression.
  • TsiR-1, TsiR-37, TsiR-26, and TsiR-34 were transfected at 50nM into two TERTp mutant (LN229 and HepG2) and two TERTp wild-type (LN18 and 293T) cell lines in 96-well plates.
  • siCTRL spiked non-targeting
  • siGABPB1 siGABPB1 were used as negative and positive control siRNAs respectively. 72 hours post transfection, the cells were harvested for RNA and changes in GABPB1L mRNA were determined by RT-qPCR analysis (ThermoFisher, Waltham, MA, Cat. No. A25600). All treatment samples were normalized to the siCTRL of their respective cell type. Error bars represent a standard error of the mean from 2 replicates. [0025] Figure 4 shows TERT expression.
  • TsiR-1, TsiR-37, TsiR-26, and TsiR-34 were transfected at 50nM into two TERTp mutant (LN229 and HepG2) and two TERTp wild-type (LN18 and 293T) cell lines in 96-well plates.
  • siCTRL scrmbled non-targeting
  • siGABPB1 were used as negative and positive control siRNAs respectively.
  • 72 hours post transfection the cells were harvested for RNA and changes in TERT mRNA were determined by RT-qPCR analysis (ThermoFisher, Waltham, MA, Cat. No. A25600). All treatment samples were normalized to the siCTRL of their respective cell type.
  • FIG. 5 shows TERT expression for specific ASOs in different cells lines.
  • TAO- 21, TAO-22, TAO-37, and TAO-40 were transfected at 25nM into two TERTp mutant (LN229 and HepG2) and two TERTp wild-type (LN18 and 293T) cell lines in 96-well plates.
  • a scrambled non-targeting LNA-ASO (Scr) was used as a negative control.
  • 72 hours post transfection the cells were harvested for RNA and changes in TERT mRNA were determined by RT-qPCR analysis (ThermoFisher, Waltham, MA, Cat. No. A25600).
  • telomerase expression may be downregulated by modulating GA Binding Protein Transcription Factor Beta 1 (GABPB1) and/or or telomerase reverse transcriptase (TERT) gene expression and/or function.
  • GABPB1 GA Binding Protein Transcription Factor Beta 1
  • TERT telomerase reverse transcriptase
  • Modulation of GABPB1 and/or TERT gene expression includes decreasing translation of a functional gene product; i.e., decreasing the translation of, and hence the levels of, GABPB1 (total or partial GABPB1 by reducing the level of GABPB1S or GABPB1L) and/or TERT.
  • compositions and methods described herein involve administering GABPB1 -hybridizing oligonucleotides to subjects having a disease, disorder and/or condition such as, but not limited to, cancer.
  • Telomerase expression is a hallmark of tumorigenesis and over 90% of human cancers aberrantly express the enzyme. Telomerase functions by elongating telomeres, the ‘TTAGGG’ DNA repeats at the end of chromosomes. The majority of normal tissues have no telomerase activity so that telomeres shorten with each successive round of cell division. Eventually, a critical telomere length is reached and cells enter replicative senescence or undergo apoptosis.
  • Telomerase reverse transcriptase is a catalytic subunit of telomerase which catalyzes the addition of nucleotides in a specific DNA sequence to the ends of a chromosome’s telomeres. This addition of repetitive DNA sequences prevents degradation of the chromosomal ends after multiple rounds of replication. Reactivation of telomerase reverse transcriptase (TERT) expression occurs in many human cancers and TERT reactivation is necessary to overcome replicative senescence (aging) and prevent apoptosis (cell death), both fundamental steps in the initiation of cancer. [0030] Thus, the GABPB1 -hybridizing oligonucleotides described herein can be administered to treat individual subjects having a disease, disorder and/or condition such as, but not limited to, cancer.
  • nucleic acid-based compositions are oligomeric polymers of at least 2 nucleoside monomers (nucleotide monomer when linked to another nucleoside. Each nucleoside monomer is further defined generally as comprising a sugar, nucleobase and backbone linker. Each of the sugar, nucleobase or backbone linker may be naturally occurring or synthetic. Certain nucleic acid-based compositions effect their outcomes through a hybridization mechanism.
  • hybridization mechanism refers to the capacity of a first oligonucleotide to form a complementary or reverse complementary (e.g., antisense) association with a second oligonucleotide via standard or non-standard hydrogen bonding.
  • the association may form along the entire length, a part, a region or intermittently along the first or second oligonucleotide.
  • nucleic acid-based compositions which are oligonucleotides that hybridize to other oligonucleotides.
  • These compositions and methods may comprise at least one antisense oligonucleotide (ASO) that downregulates the expression of the GABPB1 gene.
  • the nucleic acids are oligonucleotides or siRNAs that target GABPB1L mRNA and inhibit GABP binding to mutant TERT promoter sequences.
  • the oligonucleotides or siRNAs may target exon 9 or the intron between exon 8 and exon 9 of GABPB1L mRNA and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations.
  • TERT Telomerase reverse transcriptase
  • GABPB1 GA Binding Protein Transcription Factor Beta 1
  • the TERT gene (Genbank Acc. No. NC_000005.10; RefSeq ID: NM_198253.3 provided as SEQ ID NO: 1) encodes the catalytic subunit of telomerase and its transcriptional regulation is the rate-limiting step in telomerase activity. Certain mutations in the TERT promoter have been associated with cancer cells.
  • the mutations are typically heterozygous, occur in a mutually exclusive fashion, and both create an identical 11bp sequence. Cells harboring these mutations reactivate TERT gene expression and telomerase activity, thus becoming immortalized.
  • TERTp TERTp
  • guanine residues which may be replaced with adenine are shown below: provides the TERT promoter with G residues replaced by A for both of the hotspot mutations.
  • GABPB1 the GA binding protein transcription factor beta 1
  • GABP binds and regulates the mutant TERT promoter but not the wild-type promoter in the same cells.
  • GABP binding to the mutant TERT promoter is a common mechanism of TERT activation across multiple cancer types including glioblastoma (GBM), melanoma, hepatocellular carcinoma, and bladder cancer.
  • the present disclosure provides therapies that inhibit this mutation-specific interaction in at least the more than 50 distinct cancers that have been observed to harbor TERTp mutations to date. Many of these TERTp mutant cancers, such as GBM, have a high unmet medical need. Furthermore, contemplated herein are methods for treating cancers having TERTp mutations that have not yet been associated with this etiology.
  • GABP is a unique transcription factor within the E26 Transformation Specific (ETS) family because it must either form a heterodimer (GABPA+GABPB), or heterotetramer (two A/B dimers), in order to bind DNA and activate transcription.
  • the GABPA gene encodes a single protein isoform (Genbank Accession: NC_000021.9) that binds to DNA but cannot activate transcription.
  • GABPB1 encodes multiple protein isoforms that have a transcriptional activation domain but cannot bind to DNA. Not every isoform of GABPB1 allows for tetramer formation.
  • GABPB1L isoform
  • GABPB1L Exon 9 coding sequence GABPB1L Exon 9 coding sequence, splice sites, UTR provided as SEQ ID NO: 5
  • LZD Leucine Zipper Domain
  • At least one of the nucleotide substitutions in a GABPB1 sequence variant is conserved across multiple species. In certain embodiments, a plurality of the nucleotide substitutions in the variant are of residues that are conserved across multiple species. In certain embodiments, at least one of the nucleotide substitutions in a peptide variant is of a residue that is conserved among from human, pig, rat, mouse, dog, rabbit, cow, chicken, donkey, sheep, cat, and horse. In certain embodiments, a plurality of the nucleotide substitutions in a variant are of residues that are conserved from human, pig, rat, mouse, dog, rabbit, cow, chicken, donkey, sheep, cat, and horse.
  • nucleic acid-based compositions that “target” one or more regions and/or elements of a GABPB1 gene are designed to hybridize with the transcript of a GABPB1 gene at a specific region, site or location or at a specific element of the transcript.
  • RNA refers to any RNA transcribed from GABPB1, as the case may be depending on the context, including pre-mRNA and processed mRNA encoding a GABP protein.
  • start codon context i.e., positions -3 to -1 relative to a start codon of a gene open reading frame
  • secondary structure of an mRNA i.e., secondary structure of an mRNA, out-of-frame upstream AUGs (uAUGs), and Kozak sequence context have all been implicated in affecting mRNA stability and translation efficiency.
  • nucleic acid-based compositions described herein can decrease expression of a GABPB1 gene include alleviating the effect of downregulatory cis and trans regulators of GABPB1 translation by enhancing mRNA stability and/or translation efficiency.
  • nucleic acid-based compounds described herein target binding sites for general and sequence specific translation factors.
  • nucleic acid-based compositions described herein target sequences implicated in formation of secondary structures that may hamper scanning of the 3′ UTR by translation complexes, thereby alleviating the inhibitory effects of the secondary structures.
  • nucleic acid-based compositions described herein target sites where RNA binding proteins interact with a GABPB1 mRNA. The nucleic acid-based compositions described herein can, in some embodiments, interrupt the interaction and thereby alleviate downregulatory effects of RNA binding proteins.
  • the nucleic acid-based compositions described herein may target sequences that form secondary structures involved in RNA decay mechanisms that selectively degrade highly structured RNAs. (See Fischer, Joseph W., et al. "Structure-Mediated RNA Decay by UPF1 and G3BP1.” Molecular Cell (2020), incorporated herein by reference in its entirety.) Thus, the nucleic acid-based compositions inhibit structure-mediated RNA decay mechanisms in some embodiments, and thereby increase mRNA stability and translation efficiency. II.
  • ASOs Antisense oligonucleotides
  • ASOs are typically short (e.g., around 8 – 50 nucleotides), single-stranded oligonucleotide molecules designed to hybridize with an RNA sequence, e.g., mRNA, rRNA. Such targeting may result in either an upregulation or downregulation of the expression of the gene depending on the design of the ASO and the site of action, or localization of the ASO.
  • ASOs can target and alter messenger RNA to modulate gene expression at the level of translation. (Rinaldi, Carlo, and Matthew JA Wood.
  • ASOs modulate target gene expression through various mechanisms, including RNase H-dependent pathways, ribozyme-based RNA destruction, and steric hindrance of cellular translation machinery.
  • ASOs can also be designed to target and block critical aspects of mRNA processing, such as intron excision and polyadenylation. (Rinaldi and Wood, 2018.) Because of their capacity for broad biodistribution and the fact they can be designed to target virtually any RNA sequence, ASOs have tremendous potential as therapeutics for monogenic diseases where modulation of a single gene product’s expression can ameliorate a disease state or slow or halt disease progression.
  • DSEs downregulatory sequence elements
  • UTRs untranslated regions
  • the ASOs described herein can have an indirect downregulation effect on the RNA transcript(s) transcribed from the template strand of the target gene and/or the polypeptide(s) encoded by the target gene or mRNA.
  • the RNA transcript transcribed from the target gene is referred to as the “target transcript,” “target RNA,” “target mRNA,” or the like, as the case may be.
  • the target transcript can be an mRNA of the target gene.
  • the target transcript can exist in the cytoplasm, the nucleus, or the mitochondrion of a cell.
  • the ASOs of the present disclosure can have a downstream effect on a biological process or activity, including, for example, where targeting a first transcript affects (either by upregulating or downregulating) a second, non-target transcript.
  • the ASOs described herein are stable in rat liver microsome. In some embodiments, at least 20% of the ASOs described herein remain in rat liver microsome after 72 hours.
  • the ASOs described herein when injected intracranially into mice, do not cause weight loss or clinical signs of weakness/morbidity in mice after 1 week.
  • the nucleic acid-based compositions described herein, including ASOs described herein comprise an oligo- or polynucleotide that is at least 80% complementary to a region of the target transcript.
  • target sequence This region on the target transcript where the nucleic acid-based compositions hybridize or bind to the target transcript is referred to as the “targeted sequence” or “target site.”
  • target site This region on the target transcript where the nucleic acid-based compositions hybridize or bind to the target transcript is referred to as the “targeted sequence” or “target site.”
  • target site This region on the target transcript where the nucleic acid-based compositions hybridize or bind to the target transcript.
  • nucleic acid-based compositions described can share at least 80% sequence identity, at least 90% sequence identity, at least 95% sequence identity, at least 98% sequence identity, at least 99% sequence identity, or can be 100% identical with, the reverse complement of the targeted sequence.
  • the reverse complements of the described nucleic acid-based compositions have a high degree of sequence identity with the targeted sequence.
  • the targeted sequence can have the same length, i.e., the same number of nucleotides, as the nucleic acid based compositions or the targeted sequence can have a similar length i.e., within 1 nucleotide, within 2 nucleotides, within 3 nucleotides, within 4 nucleotides, or within 5 nucleotides compared to the length of the nucleic acid-based compositions.
  • the nucleic acid-based compositions may hybridize with all or a portion of the targeted sequence, or hybridize intermittently with the targeted sequence. In some embodiments, targeted sequence may hybridize with all or a portion of the nucleic acid-based compositions described herein, or the targeted sequence may hybridize intermittently with the nucleic acid- based compositions.
  • the targeted sequence comprises at least 8 nucleotides.
  • the targeted sequence can be 8 nucleotides, 9 nucleotides, 10 nucleotides, 11 nucleotides, 12 nucleotides, 13 nucleotides, 14 nucleotides, 15 nucleotides, 16 nucleotides, 17 nucleotides, 18 nucleotides, 19 nucleotides, 20 nucleotides, 21 nucleotides, 22 nucleotides, 23 nucleotides, 24 nucleotides, 25 nucleotides, 26 nucleotides, 27 nucleotides, 28 nucleotides, 29 nucleotides, or 30 nucleotides in length.
  • the targeted sequence is greater than 30 nucleotides in length. In some embodiments, the targeted sequence is between 6 and 18 nucleotides in length. [0053] In some embodiments, the targeted sequence is between 7 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 8 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 9 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 10 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 11 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 12 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 13 and 18 nucleotides in length.
  • the targeted sequence is between 14 and 18 nucleotides in length. In some embodiments, the targeted sequence is about 14 nucleotides in length. In some embodiments, the targeted sequence is about 15 nucleotides in length. In some embodiments, the targeted sequence is about 16 nucleotides in length. [0054]
  • the targeted sequences described herein can be any sequence derived from a GABPB1 RNA, including a pre-mRNA or an mRNA. References to sequences of GABPB1 are provided in Table 1 and is understood by those of skill in the art that such DNA may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 1.
  • the targeted sequences described herein are located in a region of GABPB1 such as, but not limited to, an exon, intron or an junction between two exons spanning between 12 nucleotides and 22 nucleotides.
  • the targeted sequence may be 14 nucleotides in length, 15 nucleotides in length, 16 nucleotides in length, 17 nucleotides in length, 18 nucleotides in length, 19 nucleotides in length, 20 nucleotides in length, 21 nucleotides in length, or 22 nucleotides in length.
  • the GABPB1- hybridizing oligonucleotides described herein can be 14-mers, 15-mers, 16-mers, 17-mers, 18-mers, 19-mers 20-mers, 21-mer, or 22-mers.
  • the targeted sequences described herein are located in exon 9 of GABPB1 or the 3’UTR of GABPB1.
  • regions or sequences that can be targeted in exon 9 of GABPB1L are described in International Patent Publication No. WO2018217975, the contents of which are herein incorporated by reference in its entirety.
  • WO2018217975 provided are regions of exon 9 or the 3' untranslated region (UTR) of GABPBIL mRNA which can be targeted by ASOs, and thus can be used to reduce TERT expression and treat cancers harboring TERT promoter mutations.
  • the targeted sequence region of GABPB1L may be, but not limited to, SEQ ID NO. 1-6 and 13-15 as described in WO2018217975, the contents of which are herein incorporated by reference.
  • ASOs which may be used to target exon 9 or the 3' UTR of GABPBIL mRNA are SEQ ID NO: 7-12 of WO2018217975, the contents of which are herein incorporated by reference in its entirety.
  • targeted sequences that target a sequence within exon 9 of GABPB1 or the intron on the exon 8-exon 9 boundary are provided in Table 2 and Table 3.
  • Nucleic acid-based compositions described herein can be designed to hybridize to a targeted sequence provided in Table 2 and Table 3.
  • Exemplary 15-mer ASOs are provided in Table 2 and SSOs are provided in Table 3.
  • * denotes a phosphorothioate bond in the backbone and + is a LNA.
  • GC content is shown as a percentage (“GCPerc”).
  • oligonucleotides are written in the 5’ to 3’ direction and as DNA oligonucleotides. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”.
  • the label “Start” refers to the position on chromosome 15 which corresponds to the start of the region of hybridization of the hybridizing oligonucleotide sequences and also to the first nucleotide of the targeted region shown in the table as the “GABP Targeted Sequence,” and “End” refers to the position on chromosome 15 which corresponds to the end of the region of hybridization.
  • Hybridization of the target sequences described herein may be at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, or 100%.
  • the GABPB1-hybridizing oligonucleotide of SEQ ID NO: 19 will hybridize to the region on chromosome 15 from Start to End of nucleotide 50570781 to 50570796.
  • SEQ ID NO: 24166 is the actual sequence from this region on chromosome 15 to which the oligonucleotide hybridizes.
  • the GABPB1-hybridizing sequence comprises SEQ ID NO: 38 or at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 38. In some embodiments, the GABPB1-hybridizing sequence comprises SEQ ID NO: 39 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 39.
  • the GABPB1 -hybridizing sequence comprises SEQ ID NO: 54 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 54.
  • the GABPBl- hybridizing sequence comprises SEQ ID NO: 57 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 57.
  • the GABPBl-hybridizing sequence comprises SEQ ID NO: 433 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 433.
  • the GABPB1 -hybrbdizing sequence comprises SEQ ID NO: 450 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 450.
  • GABPB1 15-mer Targeted Sequences and SSO Sequences is GABPB1 Hybridizing Oligonucleotides [0061]
  • the nucleic acid-based compositions described have a high degree of sequence identity with the targeted sequence of the RNA transcript. Instead of “complementary to the targeted sequence,” the nucleic acid-based compositions can also be defined as having "identity" to a region on the coding strand of the target gene. Therefore, the genomic sequence of the target gene may be used to design nucleic acid-based compositions.
  • compositions referred to herein as “GABPB1-hybridizing oligonucleotides” will thus hybridize with the targeted sequence of the RNA transcript
  • the targeted sequence may be the GABPB1L isoform, the GABPB1S isoform or may target both isoforms for a reduction in total GABPB 1.
  • Off-target hits of the nucleic acid-based compositions designed as described herein can be determined.
  • the sequences targeted by the nucleic acid-based compositions are compared against the target transcriptome represented by the human RefSeq database.
  • a number N of mismatches can be permitted in the identification of off-target hits.
  • Off-target mismatch hit number refers to the number of nucleotide mismatches permitted in an off- target hit when comparing the targeted sequence to off-target hit sequences in the transcriptome.
  • Off-target hits in the transcriptome are counted and 0 mismatch hit number and 1 mismatch hit number off-target hits are then determined.
  • M0 refers to the number of unique genes with transcripts other than the target transcripts of the nucleic acid-based composition which the nucleic acid-based composition may hybridize with or bind to with 0 mismatched base pairs.
  • M0 counts the number of unique genes among known RefSeq transcripts in the target genome, other than the target transcript, that comprise a region completely identical with the complement of the nucleic acid-based composition sequence.
  • only nucleic acid-based compositions having no off-target hits defined as no M0 hits and no Ml hits are selected for further use or testing.
  • Non-limiting examples of nucleic acid-based compositions described herein include GABPB 1 -hybridizing oligonucleotides comprising about 14 nucleotides, about 15 nucleotides, or about 16 nucleotides.
  • the sequence of GABPB1 -hybridizing oligonucleotides can have at least 60%, 70%, 80% or 90% identity to a reverse complement of a targeted sequence selected from or derived from the exon of GABPB1 mRNA or a sequence provided in Table 2 or Table 3.
  • the sequence of GABPB 1 -hybridizing oligonucleotides can have at least 60%, 70%, 80% or 90% identity to a reverse complement of a targeted sequence selected from or derived from the exon of GABPB 1 mRNA, intron of GABPB 1 mRNA or a sequence provided in Table 2, Table 3, Table 4, Table 5, Table 6, Table 7 or Table 8.
  • the nucleic acid-based compositions comprise a sequence selected from the sequences of Table 2, Table 3, Table 4, Table 5, Table 6, Table 7 or Table 8.
  • the GABPB 1 -hybridizing oligonucleotides have a 3’ tail.
  • Non-limiting examples of nucleic acid-based compositions described herein include GABPB 1 -hybridizing oligonucleotides comprising about 14 nucleotides, about 15 nucleotides, about 16 nucleotides, about 17 nucleotides, about 18 nucleotides, about 19 nucleotides, or about 20 nucleotides.
  • the sequence of GABPB 1 -hybridizing oligonucleotides can have at least 60%, 70%, 80% or 90% identity to a sequence selected from or derived from the exon of GABPB1 mRNA or a sequence provided in Table 2, Table 3, Table 4, Table 5, Table 6, Table 7, or Table 8.
  • the nucleic acid-based compositions comprise a sequence selected from the sequences of Table 2, Table 3, Table 4, Table 5, Table 6, Table 7, or Table 8.
  • the GABPB1-hybridizing oligonucleotides have a 3’ tail.
  • GABPB1-hybridizing oligonucleotides can be unmodified, or can be modified according to the modifications described herein. The GABPB1-hybridizing oligonucleotides can be fully modified. In some embodiments, GABPB1-hybridizing oligonucleotides are partially modified.
  • the nucleic acid-based compositions of the present disclosure can be produced by any suitable method, for example synthetically or by expression in cells using standard molecular biology techniques which are well-known to a person of ordinary skill in the art.
  • ASOs can be chemically synthesized or recombinantly produced using methods known in the art.
  • Antisense Oligonucleotides [0067] Some embodiments of the nucleic acid-based compositions described herein are antisense oligonucleotides (ASOs).
  • ASOs antisense oligonucleotides
  • an ASO of the present disclosure is a single-stranded oligonucleotide.
  • the ASO is a single-stranded oligodeoxynucleotide.
  • the length of an ASO determines its biodistribution upon administration.
  • the length of the ASO is optimized for biodistribution to the CNS, including but not limited to the brain.
  • the ASOs may be optimized to distribute to areas of the brain that have been effected by tumor growth as a result of glioblastoma or other brain tumors.
  • the ASO can be between about 8 and about 25 nucleotides in length.
  • an ASO of the present disclosure is at least 8, at least 9, at least 10, at least 11, 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 nucleotides in length.
  • an ASO is 8 nucleotides in length, 9 nucleotides in length, 10 nucleotides in length, 11 nucleotides in length, 12 nucleotides in length, 13 nucleotides in length, 14 nucleotides in length, 15 nucleotides in length, 16 nucleotides in length, 17 nucleotides in length, 18 nucleotides in length, 19 nucleotides in length, 20 nucleotides in length, 21 nucleotides in length, 22 nucleotides in length, 23 nucleotides in length, 24 nucleotides in length, 25 nucleotides in length or more than 25 nucleotides in length In some embodiments an ASO is up to about 30 nucleotides in length.
  • the ASO of the present disclosure has a length of about 14 nucleotides, or has a length of exactly 14 nucleotides, so as to promote biodistribution to cells of the kidney upon administration to a subject, to promote biodistribution to cells of the liver upon administration to a subject, or to promote biodistribution to cells of the kidney and cells of the liver upon administration to a subject.
  • the ASO has at least 80%, 90%, 95%, 98%, 99% or 100% complementarity to the targeted sequence.
  • the ASOs described herein have a high degree of complementarity to the targeted sequence of the RNA transcript. In some embodiments, the ASO has no more than 5, no more than 4, no more than 3, no more than 2, no more than 1, or no mismatches with reference to the targeted sequence of the RNA transcript.
  • a “strand” in the context of the present disclosure means a contiguous sequence of nucleotides, including non-naturally occurring or modified nucleotides. Two or more strands may be, or each form a part of, separate molecules, or they may be connected covalently, e.g., by a linker such as a polyethylene glycol linker.
  • An ASO duplex refers to a single ASO molecule that is at least partly self-complementary/ and capable of forming a hairpin structure, including a duplex region.
  • the term “strand” refers to one of the regions of the ASO that is complementary to another internal region of the ASO.
  • one strand will target the targeted sequence (the “guide strand”) and the other strand will act merely as a “passenger strand.”
  • the guide strand of the ASO will have no more than 5, or no more than 4 or 3, or no more than 2, or no more than 1, or no mismatches with the targeted sequence of the RNA transcript.
  • the passenger strand of a duplex ASO comprises at least one nucleotide that is not complementary/ to the corresponding nucleotide on the guide strand, called a mismatch with the guide strand.
  • the at least one mismatch with the guide strand can be at 3’ end of the passenger strand .
  • the 3’ end of the passenger strand comprises 1-5 mismatches with the guide strand.
  • the 3’ end of the passenger strand comprises 2-3 mismatches with the guide strand.
  • the 3’ end of the passenger strand comprises 6-10 mismatches with the guide strand.
  • a duplex ASO can have siRNA-like complementarity to the targeted sequence on the template strand; that is, 100% complementarity between nucleotides 2-6 from the 5’ end of the guide strand and a region of the targeted sequence.
  • Other nucleotides of the ASO can, in addition, have at least 80%, 90%, 95%, 98%, 99% or 100% complementarity to a region of the targeted sequence.
  • nucleotides 7 counted from the 5' end) until the 3' end of the ASO can have least 80%, 90%, 95%, 98%, 99% or 100% complementarity to a region of the targeted sequence.
  • small interfering RNA or “siRNA” mean a double-stranded RNA typically 20-25 nucleotides long involved in the RNA interference (RNAi) pathway and interfering with or inhibiting the expression of a specific gene.
  • the gene is the target gene of the siRNA.
  • a siRNA is usually about 21 nucleotides long, with 3' overhangs (e.g., 2 nucleotides) at each end of the two strands.
  • a siRNA inhibits target gene expression by binding to and promoting the cleavage of one or more RNA transcripts of the target gene at specific sequences.
  • the RNA transcripts are mRNA, so cleavage of mRNA results in the down-regulation of gene expression.
  • the ASO comprises a number of unpaired nucleotides at the 3' end forming 3' overhangs or tails.
  • the number of unpaired nucleotides forming the 3' overhang can range from 0 to 5 nucleotides, or 1 to 3 nucleotides, or 2 nucleotides.
  • the ASOs of the present disclosure consist of (i) a sequence having at least 80% complementarity to a targeted sequence of the target transcript; and (ii) a 3' tail of 1-5 nucleotides, which may comprise uracil residues, such as UU, UUU, or mUmU (m stands for 2’-OMe modification).
  • the ASOs of the present disclosure are duplex ASOs having a double-stranded duplex region, wherein a guide strand comprises (i) a first sequence having at least 80% complementarity to a targeted sequence; (ii) a 3' overhang of 1-5 nucleotides and a passenger strand that forms a duplex with the guide sequence. Any 3’ tail shall not be regarded as contributing to mismatches for purposes of determining complementarity between the ASO and the targeted sequence.
  • the ASOs of the present disclosure can contain a flanking sequence. The flanking sequence can be at the 3’ end or 5’ end of the ASO.
  • the flanking sequence is the sequence of a miRNA, conferring on the ASO a miRNA configuration which may be processed with Drosha and Dicer.
  • an ASO of the present disclosure has two strands and is cloned into a microRNA precursor, e.g., miR-30 backbone flanking sequence.
  • the ASOs of the present disclosure can comprise a restriction enzyme substrate or recognition sequence.
  • the restriction enzyme recognition sequence can be at the 3’ end or 5’ end of the ASO.
  • Non-limiting examples of restriction enzymes include NotI and AscI.
  • the stereochemical designation of the chiral linkage in the ASOs of the present disclosure may be controlled using methods know in the art.
  • the ASOs of the present disclosure may be chirally controlled oligonucleotides such as those described in International Patent Publication No. WO2014012081, the contents of which are herein incorporated by reference.
  • the present disclosure provides chirally controlled ASOs, and chirally controlled ASO compositions which are of high crude purity.
  • the present disclosure provides chirally controlled ASOs, and chirally controlled ASO compositions which are of high diastereomeric purity.
  • the present disclosure provides chirally controlled compositions comprising a plurality of ASOs of at least one type, wherein each type is defined by: 1) base sequence; 2) pattern of backbone linkages; 3) pattern of backbone chiral centers; and 4) pattern of backbone P-modifications.
  • the present disclosure provides chirally controlled compositions comprising a plurality of ASOs of the same type, wherein each type is defined by: 1) base sequence; 2) pattern of backbone linkages; 3) pattern of backbone chiral centers; and 4) pattern of backbone P-modifications.
  • the present disclosure provides chirally controlled compositions comprising a plurality of oligonucleotides of two or more types, wherein each type is defined by: 1) base sequence; 2) pattern of backbone linkages; 3) pattern of backbone chiral centers; and 4) pattern of backbone P-modifications.
  • the present disclosure provides ASOs comprising one or more diastereomerically pure internucleotidic linkages with respect to the chiral linkage phosphorus.
  • the present disclosure provides ASOs comprising one or more diastereomerically pure internucleotidic linkages having the structure of formula I as described in WO2014012081, the contents of which are herein incorporated by reference in their entirety.
  • the present disclosure provides ASOs comprising one or more diastereomerically pure internucleotidic linkages with respect to the chiral linkage phosphorus, and one or more phosphate diester linkages.
  • the present disclosure provides oligonucleotides comprising one or more diastereomerically pure internucleotidic linkages having the structure of formula I of WO2014012081, and one or more phosphate diester linkages.
  • the present disclosure provides ASOs comprising one or more diastereomerically pure internucleotidic linkages having the structure of formula I-c of WO2014012081, and one or more phosphate diester linkages.
  • Exemplary internucleotidic linkages including those having structures of formula I of WO2014012081, are further described below.
  • such oligonucleotides comprise a sequence further described in the application, including but not limited to those described in Tables 2 and 4, and Appendices A, B and C of WO2014012081.
  • a provided ASO comprises a combination of stereopure and stereorandom internucleotidic linkages with respect to chirality at the linkage phosphorus. For instance, in some embodiments it is desirable to have a block of one or more stereodefmed internucleotidic linkages within an ASO that is otherwise stereorandom with respect to chirality at the linkage phosphorus. In some embodiments, it is desirable to have a block of one or more internucleotidic linkages that are stereorandom within an ASO that is otherwise stereodefined with respect to chirality at the linkage phosphorus.
  • At least one nucleotide unit of a provided ASO is installed using stereoselective oligonucleotide synthesis, as described in this application, to form a pre- designed diastereomerically pure internucleotidic linkage with respect to the chiral linkage phosphorus.
  • at least two nucleotide units of a provided ASO are installed using stereoselective oligonucleotide synthesis, as described in this application, to form at least two pre-designed diastereomerically pure internucleotidic linkages with respect to the chiral linkage phosphorus.
  • At least three nucleotide units of a provided ASO are installed using stereoselective oligonucleotide synthesis, as described in WO2014012081, to form at least three pre-designed diastereomerically pure internucleotidic linkages with respect to the chiral linkage phosphorus.
  • the at least one, two, or three pre-designed diastereomerically pure internucleotidic linkages are adjacent to one another.
  • the at least one, two, or three pre-designed diastereomerically pure internucleotidic linkages are not adjacent to one another.
  • nucleotide units of a provided ASO are installed using stereoselective oligonucleotide synthesis, as described in this application, to form a pre-designed diastereomerically pure internucleotidic linkage with respect to the chiral linkage phosphorus.
  • the at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% of nucleotide units occur in one or more blocks to provide a blockmer.
  • the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different stereochemistry and/or different P-modifications relative to one another. In certain embodiments, the present disclosure provides a chirally controlled ASO, wherein at least two individual internucleotidic linkages within the ASO have different P-modifications relative to one another.
  • the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different P-modifications relative to one another, and wherein the chirally controlled ASO comprises at least one phosphate diester internucleotidic linkage.
  • the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different P-modifications relative to one another, and wherein the chirally controlled ASO comprises at least one phosphate diester internucleotidic linkage and at least one phosphorothioate diester internucleotidic linkage.
  • the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different P- modifications relative to one another, and wherein the chirally controlled ASO comprises at least one phosphorothioate triester internucleotidic linkage.
  • the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different P-modifications relative to one another, and wherein the chirally controlled ASO comprises at least one phosphate diester internucleotidic linkage and at least one phosphorothioate triester internucleotidic linkage.
  • Stereoisomers of a particular oligonucleotide can show different stability and/or activity (e.g., functional and/or toxicity properties) from each other.
  • stability and/or activity improvements may be achieved through inclusion and/or location of particular chiral structures within an oligonucleotide can be comparable to, or even better than those achieved through use of particular backbone linkages, residue modifications, etc.
  • properties (e.g., stability and/or activities) of an ASO can be adjusted by optimizing its pattern of backbone chiral centers, optionally in combination with adjustment/optimization of one or more other features (e.g., linkage pattern, nucleoside modification pattern, etc.) of the ASO as described in WO2017015555, the contents of which are herein incorporated by reference in their entirety.
  • the present disclosure provides compositions of ASOs, wherein the ASOs have a common pattern of backbone chiral centers which may enhance the stability and/or biological activity of the ASOs.
  • a pattern of backbone chiral centers provides increased stability.
  • a pattern of backbone chiral centers provides increased activity.
  • a pattern of backbone chiral centers provides increased stability and activity.
  • a pattern of backbone chiral centers such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, effectively prevents cleavage at secondary sites.
  • a pattern of backbone chiral centers such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, creates new cleavage sites.
  • a pattern of backbone chiral centers such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, minimizes the number of cleavage sites.
  • a pattern of backbone chiral centers minimizes the number of cleavage sites so that a target nucleic acid polymer is cleaved at only one site within the sequence of the target nucleic acid polymer that is complementary to the ASO.
  • a pattern of backbone chiral centers such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, enhances cleavage efficiency at a cleavage site.
  • a pattern of backbone chiral centers of the ASO improves cleavage of a target nucleic acid polymer.
  • a pattern of backbone chiral centers increases selectivity. In some embodiments, a pattern of backbone chiral centers minimizes off-target effect. In some embodiments, a pattern of backbone chiral centers increase selectivity, e.g., cleavage selectivity between two target sequences differing only by a single nucleotide polymorphism (SNP).
  • SNP single nucleotide polymorphism
  • Exemplary backbone chiral centers are provided in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, [0089]
  • the present disclosure provides compositions and methods for altering splicing of transcripts.
  • Splicing of a transcript is an essential step for the transcript to perform its biological functions in many higher eukaryotes. Defects d/ i ffi i i h li i ff bi l i l f i d/ h pathological consequences. For example, many human genetic diseases are caused by mutations that cause splicing defects, and many diseases are associated with splicing defects that are not attributed to overt mutations.
  • the present disclosure recognizes that targeting splicing, especially through compositions comprising oligonucleotides having the chemical modifications and/or stereochemistry patterns described in this disclosure, can effectively correct disease-associated aberrant splicing, and/or introduce and/or enhance beneficial splicing that lead to desired products, e.g., mRNA, proteins, etc. which can repair, restore, or add new desired biological functions.
  • desired products e.g., mRNA, proteins, etc. which can repair, restore, or add new desired biological functions.
  • desired products e.g., mRNA, proteins, etc. which can repair, restore, or add new desired biological functions.
  • desired products e.g., mRNA, proteins, etc.
  • desired products e.g., mRNA, proteins, etc.
  • desired products e.g., mRNA, proteins, etc. which can repair, restore, or add new desired biological functions.
  • compositions and methods effectively knockdown a gene; in some embodiments, such a gene is a
  • the ASOs of the present disclosure may include chemical modifications, stereochemistry and combinations thereof can be used to modulate splicing of transcripts.
  • Representative patent Publication WO2017062862 and WO2018067973 the contents of each of which are herein incorporated by reference in their entirety, are chemical modifications and patterns thereof useful for improving transcript splicing by oligonucleotides.
  • WO2017062862 and WO2018067973 describes how stereochemistry can be used to modulate transcript splicing by oligonucleotide compositions which can be used with the ASO compositions described herein.
  • combinations of chemical modifications and stereochemistry may be used to improve properties of oligonucleotides, e.g., their capabilities to alter splicing of transcripts.
  • chirally controlled oligonucleotide compositions that, when compared to a reference condition (e.g., absence of the composition, presence of a reference composition (e.g., a stereorandom composition of oligonucleotides having the same base sequence, the same chemical modifications, etc., a chirally controlled oligonucleotide composition of another stereoisomer, etc.), and combinations thereof), provide altered splicing that can deliver one or more desired biological effects, for example, increase production of desired proteins, knockdown of a gene by producing mRNA with frameshift mutations and/or premature termination codons, knockdown of a gene expressing a mRNA with a frameshift mutation and/or premature termination codon, etc.
  • the stereochemistry of an ASO described herein may be modified to improve the properties of the ASO in order to increase the effectiveness of the ASO to the target gene.
  • the stereochemistry of the ASO may be modified as described in International Patent Publication Nos. WO2018223081, WO2018223073, WO2018223056, WO2019200185, the contents of each of which are herein incorporated by reference in their entirety, where the stereochemistry of the backbone chiral centers were modified to increase the properties of the oligonucleotides to specific target genes.
  • ASOs of the present disclosure may be prepared using the stereospecific synthesis of diastereomerically pure or substantially diastereomerically pure phosphorodiamidate morpholino oligomers (PMOs) of US Patent Publication No. US20200115405, the contents of which are herein incorporated by reference, which include at least one chiral phosphorous linkage.
  • Stereochemically pure or substantially stereochemically pure monomers may be prepared by the separation of a diasteromeric mixture of monomers as described in US Patent Publication No. US20200115405, the contents of which are herein incorporated by reference, in paragraphs 48-56, and 62-102.
  • the activated monomers may be used to accomplish stereospecific coupling for the preparation of stereochemically pure dinucleotides, stereochemically pure trinucleotides, and larger stereochemically pure oligomers as described in paragraphs 58-61, and 103-161 of US Patent Publication No. US20200115405, the contents of which are herein incorporated by reference.
  • Exemplary ASO Sequences [0093] Exemplary 16-mer and 20-mer ASOs are provided in Table 4.
  • oligonucleotides are written in the 5’ to 3’ direction and as DNA oligonucleotides. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 4. GABPB116-mer and 20-mer ASO Sequences
  • Exemplary ASOs are provided in Table 5 and Table 6 which target exon 9, the intron between exon 8 and exon 9, the exon 8-exon 9 junction or total GABPB1 (shown in the table in “Target Area”). Also provided are the sequence identifiers for the target sequence for the ASOs (“Target SEQ ID”). In Table 5, “Int.E8-E9” refers to the intron between exon 8 and 9, “E9” is exon 9 and “Jun.E8-E9” is the exon 8-exon 9 junction. In Table 6, “Total” refers to targeting total GABPB1 (both GABPB1S and GABPB1L) expression. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 5. ASOs Targeting GABPB1
  • ASOs Targeting Total GABPB1 Exemplary 20-mer ASOs are provided in Table 7. Also provided are the sequence identifiers for the target sequence for the ASOs (“Target SEQ ID”).
  • Target SEQ ID is a DNA base
  • (5MdC) is a 5-Methyl-dC base
  • “Nm” is a MOE base
  • “s” is a phosphorothioate bond.
  • the oligonucleotides are written in the 5’ to 3’ direction and as DNA oligonucleotides. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 7.
  • GABPB120-mer ASO Sequences [0096] Exemplary 16-mer, 18-mer and 20-mer ASOs are provided in Table 8 (sequence identifier provided under ASO SEQ ID (with mods)) along with the identifier for the sequence without modifications (ASO SEQ ID (no mods)) and the target sequence (Target SEQ ID) .
  • dN is a DNA base
  • (5MdC)” is a 5-Methyl-dC base
  • Nm is a MOE base
  • N+ is a LNA
  • s is a phosphorothioate bond.
  • oligonucleotides are written in the 5’ to 3’ direction and as DNA oligonucleotides. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 8. 16-mer, 18-mer and 20-mer ASO Sequences
  • the ASO sequence comprises SEQ ID NO: 12335 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12335. In some embodiments, the ASO sequence comprises SEQ ID NO: 12338 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12338. In some embodiments, the ASO sequence comprises SEQ ID NO: 12363 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12363.
  • the ASO sequence comprises SEQ ID NO: 12372 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12372. In some embodiments, the ASO sequence comprises SEQ ID NO: 12376 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12376. In some embodiments, the ASO sequence comprises SEQ ID NO: 12379 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12379.
  • the ASO sequence comprises SEQ ID NO: 12382 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12382. In some embodiments, the ASO sequence comprises SEQ ID NO: 12383 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12383. In some embodiments, the ASO sequence comprises SEQ ID NO: 12384 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12384.
  • the ASO sequence comprises SEQ ID NO: 12389 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12389. In some embodiments, the ASO sequence comprises SEQ ID NO: 12393 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12393. In some embodiments, the ASO sequence comprises SEQ ID NO: 12394 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12394.
  • the ASO sequence comprises SEQ ID NO: 12395 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12395. In some embodiments, the ASO sequence comprises SEQ ID NO: 12413 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12413. In some embodiments, the ASO sequence comprises SEQ ID NO: 12421 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12421.
  • the ASO sequence comprises SEQ ID NO: 12422 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12422.
  • a selectivity index which uses the cell viability and knockdown percentages, may be used to compare a selection of ASOs and may be used as a component in determining which of the ASOs have the higher chance of success for additional studies and ultimately to be use and the compositions and methods described herein.
  • Exemplary ASO duplex Sequences [0099] Exemplary ASO duplexes of the present disclosure are provided in Table 9.
  • ASO duplexes may be used as siRNA duplexes or the individual sequences or fragments thereof may be used as siRNA sequences.
  • the target sequence and target sequence identifier (“TSID”), GC Percentage (“GC %”), the sense sequence and sense sequence identifier (“SSID”), and antisense sequence and antisense sequence identifier (“ASID”) are all shown in Table 9. Table 9.
  • ASO Duplex Sequences Bifunction Oligonucleotides [0100] Bifunctional or dual-functional nucleic acid-based compositions, including ASOs, can be designed to up-regulate the expression of a first gene and down-regulate the expression of at least one second gene.
  • nucleic acid-based compositions activate the expression of the first gene and another portion inhibits the expression of the second gene.
  • chemical Modifications [0101]
  • the terms “modification” or, as appropriate, “modified” refer to structural and/or chemical modifications with respect to A, G, T/U or C nucleotides.
  • Nucleotides in the nucleic acid-based compositions of the present disclosure can comprise non-standard nucleotides, such as non-naturally occurring nucleotides or chemically synthesized nucleotides or deoxynucleotides.
  • nucleoside is defined as a compound containing a sugar molecule (e.g., a pentose or ribose) or a derivative thereof in combination with an organic base (e.g., a purine or pyrimidine) or a derivative thereof (also referred to herein as “nucleobase”).
  • organic base e.g., a purine or pyrimidine
  • nucleotide is defined as a nucleoside including a phosphate group or other backbone linker (internucleoside linkage).
  • the nucleic acid-based compositions of the present disclosure can include any useful modification, such as to the sugar, the nucleobase, or the internucleoside linkage (e.g.
  • a pyrimidine nucleobase may be replaced or substituted with optionally substituted amino, optionally substituted thiol, optionally substituted alkyl (e.g., methyl or ethyl), or halo (e.g., chloro or fluoro).
  • modifications e.g., one or more modifications are present in each of the sugar and the internucleoside linkage.
  • Modifications according to the present disclosure may be modifications of ribonucleic acids (RNAs) to deoxyribonucleic acids (DNAs), threose nucleic acids (TNAs), glycol nucleic acids (GNAs), peptide nucleic acids (PNAs), locked nucleic acids (LNAs) or hybrids thereof.
  • the modifications include 2’-OMe-modified or 2’-O-(2- Methoxyethyl)-modified nucleotides (2’OMe and 2’MOE modifications, respectively).
  • nucleic acid-based compositions of the present disclosure can include a combination of modifications to the sugar, the nucleobase, and/or the internucleoside linkage.
  • nucleic acid-based compositions of the present disclosure may be modified with any modifications of an oligonucleotide or polynucleotide disclosed in PCT Publication WO2013/151670, the contents of which are incorporated herein by reference in their entirety, including modifications to one or more sugar, to one or more base, or to one or more internucleoside linkage of a an oligonucleotide.
  • the modified nucleotide base pairing may encompass not only the standard adenosine-thymine, adenosine-uracil, or guanosine-cytosine base pairs, but also base pairs formed between nucleotides and/or modified nucleotides comprising non-standard or modified bases, wherein the arrangement of hydrogen bond donors and hydrogen bond acceptors permits hydrogen bonding between a non-standard base and a standard base or between two complementary non-standard base structures.
  • One example of such non- standard base pairing is the base pairing between the modified nucleotide inosine and adenine, cytosine or uracil.
  • the modified nucleosides and nucleotides can include a modified nucleobase.
  • nucleobases found in RNA include, but are not limited to, adenine, guanine, cytosine, and uracil.
  • nucleobase found in DNA include, but are not limited to, adenine, guanine, cytosine, and thymine.
  • the modified nucleobase is a modified uracil.
  • nucleobases and nucleosides having a modified uracil include pseudouridine ( ⁇ ), pyridin-4- one ribonucleoside, 5-aza-uridine, 6-aza-uridine, 2-thio-5-aza-uridine, 2-thio-uridine (s 2 U), 4- thio-uridine (s 4 U), 4-thio-pseudouridine, 2-thio-pseudouridine, 5-hydroxy-uridine (ho 5 U), 5- aminoallyl-uridine, 5-halo-uridine (e.g., 5-iodo-uridine or 5-bromo-uridine), 3-methyl-uridine (m 3 U), 5-methoxy-uridine (mo 5 U), uridine 5-oxyacetic acid (cmo 5 U), uridine 5-oxyacetic acid methyl ester (mcmo 5 U), 5-carboxymethyl-uridine (cm 5 U), 1-carboxymethyl- pseudouridine, 5-carboxyhydroxymethyl-uridine (chm 5
  • the modified nucleobase is a modified cytosine.
  • exemplary nucleobases and nucleosides having a modified cytosine include 5-aza-cytidine, 6-aza- cytidine, pseudoisocytidine, 3-methyl-cytidine (m 3 C), N4-acetyl-cytidine (ac 4 C), 5-formyl- cytidine (f 5 C), N4-methyl-cytidine (m 4 C), 5-methyl-cytidine (m 5 C), 5-halo-cytidine (e.g., 5- iodo-cytidine), 5-hydroxymethyl-cytidine (hm 5 C), 1-methyl-pseudoisocytidine, pyrrolo- cytidine, pyrrolo-pseudoisocytidine, 2-thio-cytidine (s 2 C), 2-thio-5-methyl-cytidine, 4-thio- pseudo
  • the modified nucleobase is a modified adenine.
  • exemplary nucleobases and nucleosides having a modified adenine include 2-amino-purine, 2, 6- diaminopurine, 2-amino-6-halo-purine (e.g., 2-amino-6-chloro-purine), 6-halo-purine (e.g., 6- chloro-purine), 2-amino-6-methyl-purine, 8-azido-adenosine, 7-deaza-adenine, 7-deaza-8- aza-adenine, 7-deaza-2-amino-purine, 7-deaza-8-aza-2-amino-purine, 7-deaza-2,6- diaminopurine, 7-deaza-8-aza-2,6-diaminopurine, 1-methyl-adenosine (m 1 A), 2-methyl- adenine (m 2 A), N6-methyl-adenos
  • the modified nucleobase is a modified guanine.
  • exemplary nucleobases and nucleosides having a modified guanine include inosine (I), 1-methyl-inosine (m 1 I), wyosine (imG), methylwyosine (mimG), 4-demethyl-wyosine (imG-14), isowyosine (imG2), wybutosine (yW), peroxywybutosine (o2yW), hydroxywybutosine (OHyW), undermodified hydroxywybutosine (OHyW*), 7-deaza-guanosine, queuosine (Q), epoxyqueuosine (oQ), galactosyl-queuosine (galQ), mannosyl-queuosine (manQ), 7-cyano-7- deaza-guanosine (preQ0), 7-aminomethyl-7-deaza-guanosine, queuo
  • the nucleobase of the nucleotide can be independently selected from a purine, a pyrimidine, a purine or pyrimidine analog.
  • the nucleobase can each be independently selected from adenine, cytosine, guanine, uracil, or hypoxanthine.
  • the nucleobase can also include, for example, naturally-occurring and synthetic derivatives of a base, including pyrazolo[3,4-d]pyrimidines, 5-methylcytosine (5-me-C), 5- hydroxymethyl cytosine, xanthine, hypoxanthine, 2-aminoadenine, 6-methyl and other alkyl derivatives of adenine and guanine, 2-propyl and other alkyl derivatives of adenine and guanine, 2-thiouracil, 2-thiothymine and 2-thiocytosine, 5-propynyl uracil and cytosine, 6- azo uracil, cytosine and thymine, 5-uracil (pseudouracil), 4-thiouracil, 8-halo (e.g., 8-bromo), 8-amino, 8-thiol, 8-thioalkyl, 8-hydroxyl and other 8-substituted aden
  • nucleic acid-based compositions described herein comprise modifications including 5-methylcytosine (“5-me-C”) bases or 5-methylcytidine (m5c) nucleotides.
  • the nucleic acid-based compositions of the present disclosure may or may not be uniformly modified along the entire length of the molecule.
  • nucleotide e.g., purine or pyrimidine, or any one or more or all of A, G, T/U, C
  • nucleotide e.g., purine or pyrimidine, or any one or more or all of A, G, T/U, C
  • nucleotides X in a nucleic acid-based composition are modified, wherein X may be any one of nucleotides A, G, T/U, C, or any one of the combinations A+G, A+T/U, A+C, G+T/U, G+C, T/U+C, A+G+T/U, A+G+C, G+T/U+C or A+G+C.
  • X may be any one of nucleotides A, G, T/U, C, or any one of the combinations A+G, A+T/U, A+C, G+T/U, G+C, T/U+C, A+G+T/U+C or A+G+C.
  • nucleotide analogs or other modification(s) may be located at any position(s) of a nucleic acid-based composition such that the function of the nucleic acid-based composition is not substantially decreased.
  • the nucleic acid-based composition of the present disclosure may contain from about 1% to about 100% modified nucleotides (either in relation to overall nucleotide content, or in relation to one or more types of nucleotide, i.e.
  • any one or more of A, G, T/U or C) or any intervening percentage e.g., from 1% to 20%, from 1% to 25%, from 1% to 50%, from 1% to 60%, from 1% to 70%, from 1% to 80%, from 1% to 90%, from 1% to 95%, from 10% to 20%, from 10% to 25%, from 10% to 50%, from 10% to 60%, from 10% to 70%, from 10% to 80%, from 10% to 90%, from 10% to 95%, from 10% to 100%, from 20% to 25%, from 20% to 50%, from 20% to 60%, from 20% to 70%, from 20% to 80%, from 20% to 90%, from 20% to 95%, from 20% to 100%, from 50% to 60%, from 50% to 70%, from 50% to 80%, from 50% to 90%, from 50% to 95%, from 50% to 100%, from 70% to 80%, from 70% to 90%, from 70% to 95%, from 70% to 100%, from 80% to 90%, from 80% to 95%, from 90% to 100%, and from 95% to 100%
  • the nucleic acid-based compositions of the present disclosure may be modified to be a circular nucleic acid.
  • the termini of the nucleic acid- based compositions of the present disclosure may be linked by chemical reagents or enzymes, producing circular nucleic acid-based compositions that have no free ends.
  • Circular nucleic acid-based compositions are expected to be more stable than linear counterparts and to be resistant to digestion with exonucleases.
  • Circular nucleic acid-based compositions may further comprise other structural and/or chemical modifications with respect to A, G, T/U or C ribonucleotides/deoxyribonucleotides.
  • the nucleic acid-based compositions of the present disclosure may comprise a combination of modifications.
  • the nucleic acid-based compositions may comprise at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 modifications or modified nucleotides. [0119] In some embodiments, the nucleic acid-based compositions are at least 50% modified, e.g., at least 50% of the nucleotides are modified. In some embodiments, the nucleic acid-based compositions are at least 75% modified, e.g., at least 75% of the nucleotides are modified.
  • nucleic acid-based compositions are at least 10% modified in only one component of the nucleotide, with such component being the nucleobase, sugar, or linkage between nucleosides. For example, modifications may be made to at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 100% of the nucleobases, sugars, or linkages of a nucleic acid-based composition described herein.
  • RNA includes the sugar group ribose, which is a 5-membered ring having an oxygen.
  • modified nucleotides include replacement of the oxygen in ribose (e.g., with S, Se, or alkylene, such as methylene or ethylene); addition of a double bond (e.g., to replace ribose with cyclopentenyl or cyclohexenyl); ring contraction of ribose (e.g., to form a 4-membered ring of cyclobutane or oxetane); ring expansion of ribose (e.g., to form a 6- or 7-membered ring having an additional carbon or heteroatom, such as for anhydrohexitol, altritol, mannitol, cyclohexanyl, cyclohex
  • the sugar group can also contain one or more carbons that possess the opposite stereochemical configuration than that of the corresponding carbon in ribose.
  • a nucleic acid-based composition can include nucleotides containing, e.g., arabinose, as the sugar.
  • Nonlimiting examples of the sugar modification may include the modifications provided in Table 10.
  • the nucleic acid-based compositions of the present disclosure can have one or more nucleotides carrying a modification as provided in Table 10.
  • each of the nucleotides of a nucleic acid-based composition as described herein carries any one of the modifications as provided in Table 10, or none of the modifications as provided in Table 10. Table 10.
  • nucleic acid-based compositions of the present disclosure can include any modification to the internucleoside linkage (e.g.
  • modified phosphate groups include, but are not limited to, phosphorothioate, methylphosphonate, phosphoroselenates, boranophosphates, boranophosphate esters, hydrogen phosphonates, phosphoramidates, phosphorodiamidates, alkyl or aryl phosphonates, and phosphotriesters.
  • Phosphorodithioates have both non-linking oxygens replaced by sulfur.
  • the phosphate linker can also be modified by the replacement of a linking oxygen with nitrogen (bridged phosphoramidates), sulfur (bridged phosphorothioates), and carbon (bridged methylene-phosphonates).
  • the nucleic acid-based composition comprises at least one phosphorothioate linkage or methylphosphonate linkage between nucleotides.
  • the nucleic acid-based composition comprises 3’ and/or 5’ capping or overhang.
  • the nucleic acid-based composition of the present disclosure may comprise at least one inverted deoxyribonucleoside overhang (e.g., dT).
  • the inverted overhang may be at the 5’ terminus or 3’ terminus of the nucleic acid-based composition.
  • the nucleic acid-based composition of the present disclosure comprises inverted abasic modifications.
  • the inverted abasic modification(s) can be on the 5’ end, or 3’ end, or both ends of the nucleic acid-based composition.
  • the nucleic acid-based composition comprises at least one 5’-(E)-vinylphosphonate (5’-E-VP), a phosphate mimic, as a modification.
  • the nucleic acid-based composition comprises at least one glycol nucleic acid (GNA), an acyclic nucleic acid analogue, as a modification.
  • GAA glycol nucleic acid
  • the GABP1-hybridizing nucleic acid-based compositions described herein are 14-mers.
  • a nucleic acid-based composition described herein can be a 14-mer, i.e., having 14 nucleotides.
  • the 14-mers can be fully modified or partially modified according to the modifications provided herein. Configurations of varying 14-mer nucleic acid-based compositions with varying modifications can be represented as follows.
  • a 2-10-2 ASO is a 14-mer ASO having a configuration wherein two 5’ nucleotides carry a first defined modification, 10 internal nucleotides have a second defined modification, and two 3’ nucleotides carry the first defined modification, or a third modification.
  • the 14-mer ASOs described herein can have a 1-12-1 configuration, a 2-10-2 configuration, a 3-8-3 configuration, and so on.
  • the 14-mer ASOs described herein can have a 1-13 configuration, a 2-12 configuration, a 3- 11 configuration, and so on.
  • the 14-mer ASOs can have a 13-1 configuration, a 12- 2 configuration, a 11-3 configuration, and so on.
  • the 14-mer ASOs can have a 1-11-2 configuration, a 1-10-3 configuration, a 1-9-4 configuration, and so on. Similarly, the 14-mer ASOs can have a 2-11-1 configuration, a 3-10-1 configuration, a 4-9-1 configuration, and so on.
  • GABPB1-hybridizing oligonucleotides are 2-10-2 LNA-2’MOE-LNA fully modified 14-mer ASOs, wherein the two 5’ nucleotides are locked nucleic acids, the ten internal nucleotides carry 2‘-O-(2-Methoxyethyl) modifications, and the two 3’ nucleotides are locked nucleic acids.
  • the GABPB1-hybridizing nucleic acid-based compositions described herein are 15-mers.
  • a nucleic acid-based composition described herein can be a 15-mer, i.e., having 15 nucleotides.
  • the 15-mers can be fully modified or partially modified according to the modifications provided herein. Configurations of varying 15-mer nucleic acid-based compositions with varying modifications can be represented as follows.
  • a 2-11-2 ASO is a 15-mer ASO having a configuration wherein two 5’ nucleotides carry a first defined modification, 11 internal nucleotides have a second defined modification, and two 3’ nucleotides carry the first defined modification, or a third modification.
  • the 15-mer ASOs described herein can have a 1-13-1 configuration, a 2-11-2 configuration, a 3-9-3 configuration, and so on.
  • the 15-mer ASOs described herein can have a 1-14 configuration, a 2-13 configuration, a 3- 12 configuration, and so on.
  • the 15-mer ASOs can have a 14-1 configuration, a 13- 2 configuration, a 12-3 configuration, and so on.
  • the 15-mer ASOs can have a 1-12-2 configuration, a 1-11-3 configuration, a 1-10-4 configuration, and so on. Similarly, the 15-mer ASOs can have a 2-12-1 configuration, a 3-11-1 configuration, a 4-10-1 configuration, and so on.
  • GABPB1-hybridizing oligonucleotides are 2-11-2 LNA-2’MOE-LNA fully modified 15-mer ASOs, wherein the two 5’ nucleotides are locked nucleic acids, the eleven internal nucleotides carry 2‘-O-(2-Methoxyethyl) modifications, and the two 3’ nucleotides are locked nucleic acids.
  • the GABPB1-hybridizing nucleic acid-based compositions described herein are 16-mers.
  • a nucleic acid-based composition described herein can be a 16-mer, i.e., having 16 nucleotides.
  • the 16-mers can be fully modified or partially modified according to the modifications provided herein. Configurations of varying 16-mer nucleic acid-based compositions with varying modifications can be represented as follows.
  • a 2-12-2 ASO is a 16-mer ASO having a configuration wherein two 5’ nucleotides carry a first defined modification, 12 internal nucleotides have a second defined modification, and two 3’ nucleotides carry the first defined modification, or a third modification.
  • the 16-mer ASOs described herein can have a 1-14-1 configuration, a 2-12-2 configuration, a 3-10-3 configuration, and so on.
  • the 16-mer ASOs described herein can have a 1-15 configuration, a 2-14 configuration, a 3-13 configuration, and so on.
  • the 16-mer ASOs can have a 15-1 configuration, a 14-2 configuration, a 13-3 configuration, and so on.
  • the 16-mer ASOs can have a 1-13-2 configuration, a 1-12-3 configuration, a 1-11-4 configuration, and so on. Similarly, the 16-mer ASOs can have a 2-13-1 configuration, a 3- 12-1 configuration, a 4-11-1 configuration, and so on.
  • GABPB1- hybridizing oligonucleotides are 2-10-2 LNA-2’MOE-LNA fully modified 16-mer ASOs, wherein the two 5’ nucleotides are locked nucleic acids, the twelve internal nucleotides carry 2‘-O-(2-Methoxyethyl) modifications, and the two 3’ nucleotides are locked nucleic acids.
  • the GABPB1-hybridizing nucleic acid-based compositions described herein are 20-mers.
  • a nucleic acid-based composition described herein can be a 20-mer, i.e., having 20 nucleotides.
  • the 20-mers can be fully modified or partially modified according to the modifications provided herein. Configurations of varying 20-mer nucleic acid-based compositions with varying modifications can be represented as follows.
  • a 5-10-5 ASO is a 20-mer ASO having a configuration wherein five 5’ nucleotides carry a first defined modification, 10 internal nucleotides have a second defined modification, and five 3’ nucleotides carry the first defined modification, or a third modification.
  • the 20-mer ASOs described herein can have a 1-18-1 configuration, a 2-16-2 configuration, a 3-14-3 configuration, and so on.
  • the 20-mer ASOs described herein can have a 1-19 configuration, a 2-18 configuration, a 3-17 configuration, and so on.
  • the 20-mer ASOs can have a 19-1 configuration, a 18-2 configuration, a 17-3 configuration, and so on.
  • the 20-mer ASOs can have a 1-17-2 configuration, a 1-16-3 configuration, a 1-5-4 configuration, and so on. Similarly, the 20-mer ASOs can have a 2-17-1 configuration, a 3- 16-1 configuration, a 4-15-1 configuration, and so on.
  • GABP- hybridizing oligonucleotides are 5-10-5 LNA-2’MOE-LNA fully modified 20-mer ASOs, wherein the five 5’ nucleotides are locked nucleic acids, the ten internal nucleotides carry 2‘- O-(2-Methoxyethyl) modifications, and the five 3’ nucleotides are locked nucleic acids.
  • Conjugates and Combinations may result in increased stability and/or half-life and may be particularly useful in targeting the nucleic acid-based compositions of the present disclosure to specific sites in the cell, tissue, or organism.
  • the nucleic acid-based compositions of the present disclosure can be designed to be conjugated to other polynucleotides, dyes, intercalating agents (e.g. acridines), cross-linkers (e.g. psoralene, mitomycin C), porphyrins (TPPC4, texaphyrin, Sapphyrin), polycyclic aromatic hydrocarbons (e.g., phenazine, dihydrophenazine), artificial endonucleases (e.g.
  • alkylating agents phosphate, amino, mercapto, PEG (e.g., PEG-40K), MPEG, [MPEG] 2 , polyamino, alkyl, substituted alkyl, radiolabeled markers, enzymes, haptens (e.g.
  • biotin e.g., aspirin, vitamin E, folic acid
  • transport/absorption facilitators e.g., aspirin, vitamin E, folic acid
  • synthetic ribonucleases proteins, e.g., glycoproteins, or peptides, e.g., molecules having a specific affinity for a co-ligand, or antibodies e.g., an antibody, that binds to a specified cell type such as a kidney tubule cell, cancer cell, endothelial cell, or bone cell, hormones and hormone receptors, non-peptidic species, such as lipids, lectins, carbohydrates, vitamins, cofactors, or a drug.
  • a specified cell type such as a kidney tubule cell, cancer cell, endothelial cell, or bone cell
  • hormones and hormone receptors non-peptidic species, such as lipids, lectins, carbohydrates, vitamins, cofactors, or a drug.
  • nucleic acid-based compositions can be administered with, or further comprise one or more of: RNAi agents, small interfering RNAs (siRNAs), small hairpin RNAs (shRNAs), long non-coding RNAs (lncRNAs), enhancer RNAs, enhancer-derived RNAs or enhancer-driven RNAs (eRNAs), microRNAs (miRNAs), miRNA binding sites, antisense RNAs, ribozymes, catalytic DNA, tRNA, RNAs that induce triple helix formation, aptamers or vectors, and the like to achieve different functions.
  • RNAi agents small interfering RNAs
  • shRNAs small hairpin RNAs
  • lncRNAs long non-coding RNAs
  • eRNAs enhancer RNAs
  • eRNAs enhancer-derived RNAs or enhancer-driven RNAs
  • miRNAs miRNA binding sites
  • antisense RNAs ribozymes
  • RNAi agents small interfering RNAs (siRNAs), small hairpin RNAs (shRNAs), long non-coding RNAs (lncRNA), microRNAs (miRNAs), miRNA binding sites, antisense RNAs, ribozymes, catalytic DNA, tRNA, RNAs that induce triple helix formation, aptamers or vectors may comprise at least one modification or substitution.
  • the modification is a chemical substitution of the nucleic acid at a sugar position, a chemical substitution at a phosphate position, or a chemical substitution at a base position.
  • the chemical modification is incorporation of a modified nucleotide; 3′ capping; conjugation to a high molecular weight, non-immunogenic compound; conjugation to a lipophilic compound; or incorporation of phosphorothioate into the phosphate backbone.
  • the high molecular weight, non-immunogenic compound is polyalkylene glycol, or polyethylene glycol (PEG).
  • nucleic acid-based compositions of the present disclosure may be attached to a transgene so as to be co-expressed from an RNA polymerase II promoter.
  • nucleic acid-based compositions are attached to green fluorescent protein gene (GFP) or other reporter or tag.
  • GFP green fluorescent protein gene
  • nucleic acid-based compositions of the present disclosure may be attached to a DNA or RNA aptamer, thereby producing a nucleic acid-based composition-aptamer conjugate.
  • Aptamers are oligonucleotides or peptides with high selectivity, affinity, and stability. They assume specific and stable three-dimensional shapes, thereby providing highly specific, tight binding to target molecules.
  • An aptamer may be a nucleic acid species that has been engineered through repeated rounds of in vitro selection or equivalently, SELEX (systematic evolution of ligands by exponential enrichment) to bind to various molecular targets such as small molecules, proteins, nucleic acids, and even cells, tissues and organisms.
  • Nucleic acid aptamers have specific binding affinity to molecules through interactions other than classic Watson-Crick base pairing. Nucleic acid aptamers, like peptides generated by phage display or monoclonal antibodies (mAbs), are capable of specifically binding to selected targets and, through binding, block their targets’ ability to function. In some cases, aptamers may also be peptide aptamers. For any specific molecular target, nucleic acid aptamers can be identified from combinatorial libraries of nucleic acids, e.g. by SELEX. Peptide aptamers may be identified using a yeast two hybrid system.
  • a skilled person is therefore able to design suitable aptamers for delivering the nucleic acid- based compositions of the present disclosure to target cells such as kidney cells or specific sub-populations of kidney cells, like tubule cells.
  • DNA aptamers, RNA aptamers and peptide aptamers are contemplated.
  • Administration of nucleic acid-based compositions including ASOs of the present disclosure to the kidney using kidney-specific aptamers is specifically contemplated.
  • a typical nucleic acid aptamer is approximately 10-15 kDa in size (20-45 nucleotides), binds its target with at least nanomolar affinity, and discriminates against closely related targets.
  • Nucleic acid aptamers may be ribonucleic acid, deoxyribonucleic acid, or mixed ribonucleic acid and deoxyribonucleic acid. Aptamers may be single-stranded ribonucleic acid, deoxyribonucleic acid or mixed ribonucleic acid and deoxyribonucleic acid. Aptamers may comprise at least one chemical modification.
  • a suitable nucleotide length for an aptamer ranges from about 15 to about 100 nucleotides (nt), 15-30 nt, 20-25 nt, 30-100 nt, 30-60 nt, 25-70 nt, 25-60 nt, 40-60 nt, 25-40 nt, 30-40 nt, any of 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39 or 40 nt or 40-70 nt in length.
  • the sequence can be designed with sufficient flexibility such that it can accommodate interactions of aptamers with two targets at the distances described herein. Aptamers may be further modified to provide protection from nuclease and other enzymatic activities.
  • nucleic acid-based composition-aptamer conjugates may be formed using any known method for linking two moieties, such as direct chemical bond formation or linkage via a linker such as streptavidin.
  • nucleic acid-based compositions of the present disclosure may be attached to an antibody. Methods of generating antibodies against a target cell surface receptor are well known. The nucleic acid-based compositions may be attached to such antibodies with known methods, for example using RNA carrier proteins. The resulting complex may then be administered to a subject and taken up by the target cells via receptor- mediated endocytosis.
  • nucleic acid-based compositions of the present disclosure may be conjugated with lipid moieties such as a cholesterol moiety (Letsinger et al., Proc. Natl. Acid. Sci. USA, 1989, 86: 6553-6556), cholic acid (Manoharan et al., Biorg. Med. Chem. Let., 1994, 4:1053-1060), a thioether, e.g., beryl-5-tritylthiol (Manoharan et al., Ann. N.Y. Acad. Sci., 1992, 660:306-309; Manoharan et al., Biorg. Med. Chem.
  • lipid moieties such as a cholesterol moiety (Letsinger et al., Proc. Natl. Acid. Sci. USA, 1989, 86: 6553-6556), cholic acid (Manoharan et al., Biorg. Med. Chem. Let., 1994, 4:1053-1060),
  • 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 (Manoharan et al., Tetrahedron Lett., 1995, 36:3651-3654), a palmityl moiety (Mishra et al., Biochim. Biophys. Acta, 1995, 1264:229-237), or an octadecylamine or hexylamino-carbonyloxycholesterol moiety (Crooke et al., J. Pharmacol. Exp.
  • the nucleic acid-based compositions of the present disclosure are conjugated with a ligand.
  • the ligand may be any ligand disclosed in US 20130184328 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety.
  • the conjugate has a formula of Ligand- [linker] optional -[tether] optional -oligonucleotide agent.
  • the oligonucleotide agent may comprise a subunit having formula (I) disclosed by US 20130184328.
  • the ligand may be any ligand disclosed in US 20130317081 to Akinc et al., the contents of which are incorporated herein by reference in their entirety, such as a lipid, a protein, a hormone, or a carbohydrate ligand of Formula II-XXVI.
  • the ligand may be coupled with the nucleic acid-based composition with a bivalent or trivalent branched linker in Formula XXXI-XXXV disclosed in Akinc.
  • Representative U.S. patents that teach the preparation of such nucleic acid/lipid conjugates include, but are not limited to, U.S. Pat. Nos.
  • nucleic acid-based compositions of the present disclosure may be provided in combination with other active ingredients known to have an effect in the particular method being considered.
  • the other active ingredients may be administered simultaneously, separately, or sequentially with the nucleic acid-based compositions.
  • nucleic acid-based compositions of the present disclosure are administered with nucleic acid- based compositions, ASOs, or other agents modulating a different target gene.
  • the nucleic acid-based compositions are conjugated with a carbohydrate ligand, such as any carbohydrate ligand disclosed in US Pat No. 8106022 and 8828956 to Manoharan et al.
  • the carbohydrate ligand may be monosaccharide, disaccharide, trisaccharide, tetrasaccharide, oligosaccharide, or polysaccharide.
  • carbohydrate-conjugated RNA agents may target a particular cell or tissue type.
  • the nucleic acid-based compositions are conjugated with more than one carbohydrate ligand, preferably two or three. In some embodiments, the nucleic acid-based compositions are conjugated with one or more galactose moiety.
  • nucleic acid-based compositions are conjugated with at least one (e.g., two or three or more) lactose molecules (lactose is a glucose coupled to a galactose). In some embodiments, the nucleic acid-based compositions are conjugated with at least one (e.g., two or three or more) N-Acetyl-Galactosamine (GalNAc), N-Ac-Glucosamine (GluNAc), or mannose (e.g., mannose-6-phosphate). [0147] In some embodiments, nucleic acid-based compositions of the present disclosure are administered with a small interfering RNA or siRNA that inhibits the expression of a gene.
  • lactose is a glucose coupled to a galactose
  • nucleic acid-based compositions are conjugated with at least one (e.g., two or three or more) N-Acetyl-Galactosamine (GalNAc), N-Ac-Glucosamine
  • nucleic acid-based compositions of the present disclosure are administered with one or more drugs for therapeutic purposes, including for treatment of ADPKD.
  • compositions comprising at least one pharmaceutically acceptable carrier and a nucleic acid-based composition, i.e., oligo- or polynucleotide molecule, including in some instances an antisense oligonucleotide (ASO) that downregulates a target gene.
  • ASO antisense oligonucleotides
  • expression constructs may be operably linked to one or more regulatory nucleotide sequences in an expression construct.
  • the one or more regulatory nucleotide sequences may include, but are not limited to, promoter sequences, leader or signal sequences, transcriptional start and termination sequences, and enhancer or activator sequences. Constitutive or inducible promoters as known in the art are also contemplated.
  • the promoters may be either naturally occurring promoters, or hybrid promoters that combine elements of more than one promoter.
  • An expression construct may be present in a cell on an episome, such as a plasmid, or the expression construct may be inserted in a chromosome.
  • the expression vector includes a selectable marker gene to allow the selection of transformed host cells.
  • Certain embodiments include an expression vector encoding a GABPB1-hybridizing oligonucleotide sequence operably linked to at least one regulatory sequence. Regulatory sequences for use herein include promoters, enhancers, and other expression control elements.
  • an expression vector is designed considering the choice of the host cell to be transformed, the particular GABP1L sequence to be expressed, the vector's copy number, the ability to control that copy number, or the expression of other proteins encoded by the vector, such as antibiotic markers.
  • the gene products of the combinatorial libraries generated by the combinatorial mutagenesis of the nucleic acids described herein may be screened.
  • Such screening methods include, for example, cloning the gene library into replicable expression vectors, transforming appropriate cells with the resulting library of vectors, and expressing the combinatorial genes under conditions to form such library.
  • the screening methods optionally further comprise detecting a desired activity and isolating a product detected.
  • Each of the illustrative assays described below are amenable to high-throughput analysis as necessary to screen large numbers of degenerate sequences created by combinatorial mutagenesis techniques.
  • the nucleic acids described herein may be expressed in microorganisms.
  • the nucleic acid may be expressed in a bacterial system, for example, in Bacillus brevis, Bacillus megaterium, Bacillus subtilis, Caulobacter crescentus, Escherichia coli and their derivatives.
  • Exemplary promoters include the l- arabinose inducible araBAD promoter (PBAD), the lac promoter, the l-rhamnose inducible rhaP BAD promoter, the T7 RNA polymerase promoter, the trc and tac promoter, the lambda phage promoter Pl, and the anhydrotetracycline-inducible tetA promoter/operator.
  • the nucleic acids described herein may be expressed in a yeast expression system.
  • promoters which may be used in yeast vectors include the promoters for 3-phosphoglycerate kinase (Hitzeman et al., J. Biol. Chem. 255:2073 (1980)); other glycolytic enzymes (Hess et al., J. Adv. Enzyme Res. 7:149 (1968); Holland et al., Biochemistry 17:4900 (1978).
  • Others promoters are from, e.g., enolase, glyceraldehyde-3-phosphate dehydrogenase, hexokinase, pyruvate decarboxylase, phosphofructokinase, glucose-6-phosphate isomerase, 3-phosphoglycerate mutase, pyruvate kinase, triosephosphate isomerase, phosphoglucose isomerase, glucokinase alcohol oxidase I (AOX1), alcohol dehydrogenase 2, isocytochrome C, acid phosphatase, degradative enzymes associated with nitrogen metabolism, and the aforementioned glyceraldehyde-3-phosphate dehydrogenase, and enzymes responsible for maltose and galactose utilization.
  • enolase e.g., enolase, glyceraldehyde-3-phosphate dehydrogenase, hexokin
  • Any plasmid vector containing a yeast-compatible promoter and termination sequences, with or without an origin of replication, is suitable.
  • Certain yeast expression systems are commercially available, for example, from Clontech Laboratories, Inc. (Palo Alto, Calif., e.g. Pyex 4T family of vectors for S. cerevisiae), Invitrogen (Carlsbad, Calif., e.g. Ppicz series Easy Select Pichia Expression Kit) and Stratagene (La Jolla, Calif., e.g. ESP.TM. Yeast Protein Expression and Purification System for S. pombe and Pesc vectors for S. cerevisiae).
  • the nucleic acids described herein may be expressed in mammalian expression systems.
  • mammalian promoters include, for example, promoters from the following genes: ubiquitin/S27a promoter of the hamster (WO 97/15664), Simian vacuolating virus 40 (SV40) early promoter, adenovirus major late promoter, mouse metallothionein-I promoter, the long terminal repeat region of Rous Sarcoma Virus (RSV), mouse mammary tumor virus promoter (MMTV), Moloney murine leukemia virus Long Terminal repeat region, and the early promoter of human Cytomegalovirus (CMV).
  • ubiquitin/S27a promoter of the hamster WO 97/15664
  • Simian vacuolating virus 40 (SV40) early promoter adenovirus major late promoter
  • mouse metallothionein-I promoter the long terminal repeat region of Rous Sarcoma Virus (RSV)
  • MMTV mouse mamm
  • heterologous mammalian promoters examples include the actin, immunoglobulin or heat shock promoter(s).
  • a yeast alcohol oxidase promoter is used.
  • promoters for use in mammalian host cells can be obtained from the genomes of viruses such as polyoma virus, fowlpox virus (UK 2,211,504 published 5 Jul. 1989), bovine papilloma virus, avian sarcoma virus, cytomegalovirus, a retrovirus, hepatitis-B virus and Simian Virus 40 (SV40).
  • viruses such as polyoma virus, fowlpox virus (UK 2,211,504 published 5 Jul. 1989), bovine papilloma virus, avian sarcoma virus, cytomegalovirus, a retrovirus, hepatitis-B virus and Simian Virus 40 (SV40).
  • viruses such as polyoma virus, fowlpox virus (UK 2,211,504 published
  • nucleic acids described herein may be expressed in insect cell expression systems.
  • Eukaryotic expression systems employing insect cell hosts may rely on either plasmid or baculoviral expression systems.
  • Typical insect host cells are derived from the fall army worm (Spodoptera frugiperda). For expression of a foreign protein these cells are infected with a recombinant form of the baculovirus Autographa californica nuclear polyhedrosis virus which has the gene of interest expressed under the control of the viral polyhedron promoter.
  • Other insects infected by this virus include a cell line known commercially as "High 5" (Invitrogen) which is derived from the cabbage looper (Trichoplusia ni).
  • Baculovirus sometimes used is the Bombyx mori nuclear polyhedorsis virus which infect the silkworm (Bombyx mori).
  • Numerous baculovirus expression systems are commercially available, for example, from Thermo Fisher (Bac-N- Blue TM k or BAC-TO-BAC TM Systems), Clontech (BacPAK TM Baculovirus Expression System), Novagen (Bac Vector System TM ), or others from Pharmingen or Quantum Biotechnologies.
  • Another insect cell host is the common fruit fly, Drosophila melanogaster, for which a transient or stable plasmid based transfection kit is offered commercially by Thermo Fisher (The DES TM System).
  • cells are transformed with vectors that express a nucleic acid described herein. Transformation techniques for inserting new genetic material into eukaryotic cells, including animal and plant cells, are well known. Viral vectors may be used for inserting expression cassettes into host cell genomes. Alternatively, the vectors may be transfected into the host cells. Transfection may be accomplished by methods as described in the art such as, but not limited to, calcium phosphate precipitation, electroporation, optical transfection, protoplast fusion, impalefection, and hydrodynamic delivery.
  • compositions may additionally comprise a pharmaceutically acceptable excipient, which, as used herein, includes, but is not limited to, any and all solvents, dispersion media, diluents, or other liquid vehicles, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, and the like, as suited to the particular dosage form desired.
  • a pharmaceutically acceptable excipient includes, but is not limited to, any and all solvents, dispersion media, diluents, or other liquid vehicles, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, and the like, as suited to the particular dosage form desired.
  • excipients for formulating pharmaceutical compositions and techniques for preparing the composition are known in the art (see Remington: The Science and Practice of Pharmacy, 21 st Edition, A. R. Gennaro, Lippincott, Williams & Wilkins, Baltimore, MD,
  • compositions are administered to humans, human patients or subjects.
  • active ingredient generally refers to a nucleic acid-based composition to be delivered as described herein.
  • compositions are principally directed to pharmaceutical compositions which are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to any other animal, e.g., to non-human animals, e.g. non-human mammals. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and/or perform such modification with merely ordinary, if any, experimentation.
  • compositions described herein may be prepared by any method known or hereafter developed in the art of pharmacology. In general, such preparatory methods include the step of bringing the active ingredient into association with an excipient and/or one or more other accessory ingredients, and then, if necessary and/or desirable, dividing, shaping and/or packaging the product into a desired single- or multi-dose unit.
  • a pharmaceutical composition in accordance with the disclosure may be prepared, packaged, and/or sold in bulk, as a single unit dose, and/or as a plurality of single unit doses.
  • a “unit dose” is discrete amount of the pharmaceutical composition comprising a predetermined amount of the active ingredient.
  • the amount of the active ingredient is generally equal to the dosage of the active ingredient which would be administered to a subject and/or a convenient fraction of such a dosage such as, for example, one-half or one-third of such a dosage.
  • compositions described herein may contain at least one nucleic acid-based composition, e.g., ASO.
  • the formulations may contain 1, 2, 3, 4 or 5 nucleic acid-based compositions with different sequences.
  • the formulation contains at least three nucleic acid-based compositions with different sequences. In some embodiments, the formulation contains at least five nucleic acid- based compositions with different sequences.
  • the nucleic acid-based compositions of the present disclosure can be formulated using one or more excipients to: (1) increase stability; (2) increase cell transfection; (3) permit the sustained or delayed release (e.g., from a depot formulation of the nucleic acid- based composition); (4) alter the biodistribution (e.g., target the nucleic acid-based composition to specific tissues or cell types); (5) increase the translation of encoded protein in vivo; and/or (6) alter the release profile of encoded protein in vivo.
  • excipients of the present disclosure can include, without limitation, lipidoids, liposomes, lipid nanoparticles, polymers, lipoplexes, core-shell nanoparticles, peptides, proteins, cells transfected with nucleic acid- based compositions (e.g., for transplantation into a subject), hyaluronidase, nanoparticle mimics and combinations thereof.
  • the formulations of the disclosure can include one or more excipients, each in an amount that together increases the stability of the nucleic acid-based compositions and/or increases cell transfection by the nucleic acid-based compositions.
  • the nucleic acid-based compositions of the present disclosure may be formulated using self-assembled nucleic acid nanoparticles.
  • Pharmaceutically acceptable carriers, excipients, and delivery agents for nucleic acids that may be used in the formulation with the nucleic acid-based compositions of the present disclosure are disclosed in International Publication WO 2013/090648, the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-based compositions of the present disclosure comprise two single RNA strands that are annealed to form a double-stranded oligonucleotide as the active ingredient.
  • the composition further comprises a salt buffer composed of 50mM Tris-HCl, pH 8.0, 100mM NaCl and 5mM EDTA.
  • the nucleic acid-based compositions of the present disclosure may be delivered with dendrimers. Dendrimers are highly branched macromolecules.
  • the nucleic acid-based compositions are complexed with structurally flexible poly(amidoamine) (PAMAM) dendrimers for targeted in vivo delivery.
  • PAMAM structurally flexible poly(amidoamine)
  • the complex is called a nucleic acid-based composition-dendrimer complex.
  • Dendrimers have a high degree of molecular uniformity, narrow molecular weight distribution, specific size and shape characteristics, and a highly-functionalized terminal surface.
  • the manufacturing process is a series of repetitive steps starting with a central initiator core. Each subsequent growth step represents a new generation of polymers with a larger molecular diameter and molecular weight, and more reactive surface sites than the preceding generation.
  • PAMAM dendrimers are efficient nucleotide delivery systems that bear primary amine groups on their surface and also a tertiary amine group inside of the structure.
  • the primary amine group participates in nucleotide binding and promotes their cellular uptake, while the buried tertiary amino groups act as a proton sponge in endosomes and enhance the release of nucleic acid into the cytoplasm.
  • These dendrimers protect the nucleic acid-based composition carried by them from ribonuclease degradation and achieves substantial release of nucleic acid-based composition over an extended period of time via endocytosis for efficient gene targeting.
  • the in vivo efficacy of these nanoparticles have previously been evaluated where biodistribution studies show that the dendrimers preferentially accumulate in peripheral blood mononuclear cells and live with no discernible toxicity (see Zhou et al., Molecular Ther. 2011 Vol.
  • PAMAM dendrimers may comprise a triethanolamine (TEA) core, a diaminobutane (DAB) core, a cystamine core, a diaminohexane (HEX) core, a diaminododecane (DODE) core, or an ethylenediamine (EDA) core.
  • TEA triethanolamine
  • DAB diaminobutane
  • HEX diaminohexane
  • DODE diaminododecane
  • EDA ethylenediamine
  • PAMAM dendrimers comprise a TEA core or a DAB core.
  • Lipidoids [0170] The synthesis of lipidoids has been extensively described and formulations containing these compounds are particularly suited for delivery of oligonucleotides or nucleic acids (see Mahon et al., Bioconjug Chem. 201021:1448-1454; Schroeder et al., J Intern Med. 2010267:9-21; Akinc et al., Nat Biotechnol. 200826:561-569; Love et al., Proc Natl Acad Sci U S A. 2010107:1864-1869; Siegwart et al., Proc Natl Acad Sci U S A. 2011108:12996- 3001; all of which are incorporated herein in their entireties).
  • nucleic acid-based compositions can be administered by various means including, but not limited to, intravenous (IV), intramuscular (IM), subcutaneous (SC), intraparenchymal (IPa), intrathecal (IT), or intracerebroventricular (ICV) administration.
  • IV intravenous
  • IM intramuscular
  • SC subcutaneous
  • IPa intraparenchymal
  • IMV intrathecal
  • ICV intracerebroventricular
  • nucleic acids may be affected by many parameters, including, but not limited to, the formulation composition, nature of particle PEGylation, degree of loading, oligonucleotide to lipid ratio, and biophysical parameters such as, but not limited to, particle size (Akinc et al., Mol Ther. 200917:872-879; the contents of which are herein incorporated by reference in its entirety).
  • particle size Akinc et al., Mol Ther. 200917:872-879; the contents of which are herein incorporated by reference in its entirety.
  • small changes in the anchor chain length of poly(ethylene glycol) (PEG) lipids may result in significant effects on in vivo efficacy.
  • Formulations with the different lipidoids including, but not limited to penta[3-(1- laurylaminopropionyl)]-triethylenetetramine hydrochloride (TETA–5LAP; aka 98N12-5, see Murugaiah et al., Analytical Biochemistry, 401:61 (2010); the contents of which are herein incorporated by reference in its entirety), C12-200 (including derivatives and variants), and MD1, can be tested for in vivo activity.
  • TETA–5LAP penta[3-(1- laurylaminopropionyl)]-triethylenetetramine hydrochloride
  • 98N12-5 see Murugaiah et al., Analytical Biochemistry, 401:61 (2010); the contents of which are herein incorporated by reference in its entirety
  • C12-200 including derivatives and variants
  • MD1 can be tested for in vivo activity.
  • the lipidoid referred to herein as “C12-200” is disclosed by Love et al., Proc Natl Acad Sci U S A. 2010107:1864-1869 and Liu and Huang, Molecular Therapy. 2010669- 670; the contents of both of which are herein incorporated by reference in their entirety.
  • the lipidoid formulations can include particles comprising either 3 or 4 or more components in addition to the nucleic acid-based compositions.
  • formulations with certain lipidoids include, but are not limited to, 98N12-5 and may contain 42% lipidoid, 48% cholesterol and 10% PEG (C14 alkyl chain length).
  • formulations with certain lipidoids include, but are not limited to, C12-200 and may contain 50% lipidoid, 10% disteroylphosphatidyl choline, 38.5% cholesterol, and 1.5% PEG-DMG.
  • nucleic acid-based compositions formulated with a lipidoid for systemic intravenous administration include, but are not limited to, C12-200 and may contain 50% lipidoid, 10% disteroylphosphatidyl choline, 38.5% cholesterol, and 1.5% PEG-DMG.
  • a final optimized intravenous formulation using nucleic acid-based compositions and comprising a lipid molar composition of 42% 98N12-5, 48% cholesterol, and 10% PEG-lipid with a final weight ratio of about 7.5 to 1 total lipid to nucleic acid-based compositions and a C14 alkyl chain length on the PEG lipid, with a mean particle size of roughly 50–60 nm, can result in the distribution of the formulation to be greater than 90% to the liver.
  • an intravenous formulation using a C12-200 may have a molar ratio of 50/10/38.5/1.5 of C12-200/disteroylphosphatidyl choline/cholesterol/PEG-DMG, with a weight ratio of 7 to 1 total lipid to nucleic acid and a mean particle size of 80 nm may be effective to deliver nucleic acid-based compositions (see, Love et al., Proc Natl Acad Sci U S A. 2010107:1864-1869, the contents of which are herein incorporated by reference in its entirety).
  • an MD1 lipidoid-containing formulation may be used to effectively deliver nucleic acid-based compositions to hepatocytes in vivo.
  • the characteristics of optimized lipidoid formulations for intramuscular or subcutaneous routes may vary significantly depending on the target cell type and the ability of formulations to diffuse through the extracellular matrix into the blood stream. While a particle size of less than 150 nm may be desired for effective hepatocyte delivery due to the size of the endothelial fenestrae (see, Akinc et al., Mol Ther.
  • lipidoid-formulated nucleic acid-based compositions to deliver the formulation to other cells types including, but not limited to, endothelial cells, myeloid cells, and muscle cells may not be similarly size-limited.
  • Use of lipidoid formulations to deliver siRNA in vivo to other non-hepatocyte cells such as myeloid cells and endothelium has been reported (see Akinc et al., Nat Biotechnol. 200826:561-569; Leuschner et al., Nat Biotechnol. 201129:1005-1010; Cho et al. Adv. Funct. Mater.
  • lipidoid formulations may have a similar component molar ratio. Different ratios of lipidoids and other components including, but not limited to, disteroylphosphatidyl choline, cholesterol and PEG-DMG, may be used to optimize the formulation of nucleic acid-based compositions for delivery to different cell types including, but not limited to, hepatocytes, myeloid cells, muscle cells, etc.
  • the component molar ratio may include, but is not limited to, 50% C12-200, 10% disteroylphosphatidyl choline, 38.5% cholesterol, and %1.5 PEG-DMG (see Leuschner et al., Nat Biotechnol 201129:1005-1010; the contents of which are herein incorporated by reference in its entirety).
  • the use of lipidoid formulations for the localized delivery of nucleic acids to cells via either subcutaneous or intramuscular delivery may not require all of the formulation components desired for systemic delivery, and as such may comprise only the lipidoid and nucleic acid-based compositions.
  • nucleic acid-based compositions of the disclosure can be formulated using one or more liposomes, lipoplexes, or lipid nanoparticles.
  • pharmaceutical compositions of nucleic acid-based compositions include liposomes. Liposomes are artificially-prepared vesicles which may primarily be composed of a lipid bilayer and may be used as a delivery vehicle for the administration of nutrients and pharmaceutical formulations.
  • Liposomes can be of different sizes such as, but not limited to, a multilamellar vesicle (MLV) which may be hundreds of nanometers in diameter and may contain a series of concentric bilayers separated by narrow aqueous compartments, a small unicellular vesicle (SUV) which may be smaller than 50 nm in diameter, and a large unilamellar vesicle (LUV) which may be between 50 and 500 nm in diameter.
  • MLV multilamellar vesicle
  • SUV small unicellular vesicle
  • LUV large unilamellar vesicle
  • Liposome design may include, but is not limited to, opsonins or ligands in order to improve the attachment of liposomes to unhealthy tissue or to activate events such as, but not limited to, endocytosis.
  • Liposomes may contain a low or a high pH in order to improve the delivery of the pharmaceutical formulations.
  • the formation of liposomes may depend on the physicochemical characteristics such as, but not limited to, the pharmaceutical formulation entrapped and the liposomal ingredients, the nature of the medium in which the lipid vesicles are dispersed, the effective concentration of the entrapped substance and its potential toxicity, any additional processes involved during the application and/or delivery of the vesicles, the optimization size, polydispersity and the shelf-life of the vesicles for the intended application, and the batch-to- batch reproducibility and possibility of large-scale production of safe and efficient liposomal products.
  • compositions described herein may include, without limitation, liposomes such as those formed from 1,2-dioleyloxy-N,N- dimethylaminopropane (DODMA) liposomes, DiLa2 liposomes from Marina Biotech (Bothell, WA), 1,2-dilinoleyloxy-3-dimethylaminopropane (DLin-DMA), 2,2-dilinoleyl-4-(2- dimethylaminoethyl)-[1,3]-dioxolane (DLin-KC2-DMA), and MC3 (US20100324120; the contents of which are herein incorporated by reference in its entirety) and liposomes which may deliver small molecule drugs such as, but not limited to, DOXIL® from Janssen Biotech, Inc.
  • DODMA 1,2-dioleyloxy-N,N- dimethylaminopropane
  • DLin-DMA 1,2-dilinoleyloxy-3-dimethylaminopropane
  • compositions described herein may include, without limitation, liposomes such as those formed from the synthesis of stabilized plasmid- lipid particles (SPLP) or stabilized nucleic acid lipid particle (SNALP) that have been previously described and shown to be suitable for oligonucleotide delivery in vitro and in vivo (see Wheeler et al. Gene Therapy. 19996:271-281; Zhang et al. Gene Therapy. 1999 6:1438-1447; Jeffs et al. Pharm Res. 200522:362-372; Morrissey et al., Nat Biotechnol. 2005 2:1002-1007; Zimmermann et al., Nature.
  • liposomes such as those formed from the synthesis of stabilized plasmid- lipid particles (SPLP) or stabilized nucleic acid lipid particle (SNALP) that have been previously described and shown to be suitable for oligonucleotide delivery in vitro and in vivo (see Wheeler et al. Gene Therapy.
  • the liposome formulations may be composed of 3 to 4 lipid components in addition to the nucleic acid-based compositions.
  • a liposome can contain, but is not limited to, 55% cholesterol, 20% disteroylphosphatidyl choline (DSPC), 10% PEG-S-DSG, and 15% 1,2-dioleyloxy-N,N- dimethylaminopropane (DODMA), as described by Jeffs et al.
  • DSPC disteroylphosphatidyl choline
  • PEG-S-DSG 10% PEG-S-DSG
  • DODMA 1,2-dioleyloxy-N,N- dimethylaminopropane
  • certain liposome formulations may contain, but are not limited to, 48% cholesterol, 20% DSPC, 2% PEG-c-DMA, and 30% cationic lipid, where the cationic lipid can be 1,2-distearloxy-N,N- dimethylaminopropane (DSDMA), DODMA, DLin-DMA, or 1,2-dilinolenyloxy-3- dimethylaminopropane (DLenDMA), as described by Heyes et al.
  • DSDMA 1,2-distearloxy-N,N- dimethylaminopropane
  • DODMA 1,2-dilinolenyloxy-3- dimethylaminopropane
  • the nucleic acid-lipid particle may comprise a cationic lipid comprising from about 50 mol % to about 85 mol % of the total lipid present in the particle; a non-cationic lipid comprising from about 13 mol % to about 49.5 mol % of the total lipid present in the particle; and a conjugated lipid that inhibits aggregation of particles comprising from about 0.5 mol % to about 2 mol % of the total lipid present in the particle as described in WO2009127060 to Maclachlan et al, the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-lipid particle may be any nucleic acid-lipid particle disclosed in US2006008910 to Maclachlan et al., the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-lipid particle may comprise a cationic lipid of Formula I, a non-cationic lipid, and a conjugated lipid that inhibits aggregation of particles.
  • the nucleic acid-based compositions of the present disclosure may be formulated in a lipid vesicle which may have crosslinks between functionalized lipid bilayers.
  • the liposome may contain a sugar-modified lipid disclosed in US5595756 to Bally et al., the contents of which are incorporated herein by reference in their entirety.
  • the lipid may be a ganglioside and cerebroside in an amount of about 10 mol percent.
  • the nucleic acid-based compositions of the present disclosure may be formulated in a liposome comprising a cationic lipid.
  • the liposome may have a molar ratio of nitrogen atoms in the cationic lipid to the phosphates in the nucleic acid-based compositions (N:P ratio) of between 1:1 and 20:1 as described in International Publication No.
  • the liposome may have a N:P ratio of greater than 20:1 or less than 1:1.
  • the nucleic acid-based compositions of the present disclosure may be formulated in a lipid-polycation complex. The formation of the lipid- polycation complex may be accomplished by methods known in the art and/or as described in U.S. Pub. No. 20120178702, the contents of which are herein incorporated by reference in its entirety.
  • the polycation may include a cationic peptide or a polypeptide such as, but not limited to, polylysine, polyornithine and/or polyarginine and the cationic peptides described in International Pub. No. WO2012013326; herein incorporated by reference in its entirety.
  • the nucleic acid-based compositions may be formulated in a lipid-polycation complex which may further include a neutral lipid such as, but not limited to, cholesterol or dioleoyl phosphatidylethanolamine (DOPE).
  • DOPE dioleoyl phosphatidylethanolamine
  • the liposome formulation may be influenced by, but not limited to, the selection of the cationic lipid component, the degree of cationic lipid saturation, the nature of the PEGylation, ratio of all components and biophysical parameters such as size.
  • the liposome formulation was composed of 57.1 % cationic lipid, 7.1% dipalmitoylphosphatidylcholine, 34.3 % cholesterol, and 1.4% PEG-c-DMA.
  • the ratio of PEG in the lipid nanoparticle (LNP) formulations may be increased or decreased and/or the carbon chain length of the PEG lipid may be modified from C14 to C18 to alter the pharmacokinetics and/or biodistribution of the LNP formulations.
  • LNP formulations may contain 1-5% of the lipid molar ratio of PEG-c-DOMG as compared to the cationic lipid, DSPC and cholesterol.
  • the PEG-c-DOMG may be replaced with a PEG lipid such as, but not limited to, PEG-DSG (1,2-Distearoyl-sn-glycerol, methoxypolyethylene glycol) or PEG-DPG (1,2-Dipalmitoyl-sn-glycerol, methoxypolyethylene glycol).
  • PEG-DSG 1,2-Distearoyl-sn-glycerol, methoxypolyethylene glycol
  • PEG-DPG 1,2-Dipalmitoyl-sn-glycerol, methoxypolyethylene glycol
  • the cationic lipid may be selected from any lipid known in the art such as, but not limited to, DLin-MC3-DMA, DLin- DMA, C12-200 and DLin-KC2-DMA.
  • the nucleic acid-based compositions of the present disclosure may be formulated in a lipid nanoparticle such as the lipid nanoparticles described in International Publication No. WO2012170930, the contents of which are herein incorporated by reference in its entirety.
  • the cationic lipid which may be used in formulations of the present disclosure may be selected from, but not limited to, a cationic lipid described in International Publication Nos.
  • the cationic lipid may be selected from, but not limited to, formula A described in International Publication Nos.
  • the cationic lipid may be selected from, but not limited to, formula CLI-CLXXIX of International Publication No. WO2008103276, formula CLI-CLXXIX of US Patent No. 7,893,302, formula CLI-CLXXXXII of US Patent No. 7,404,969 and formula I-VI of US Patent Publication No.
  • the cationic lipid may be a multivalent cationic lipid such as the cationic lipid disclosed in US Patent No. 7223887 to Gaucheron et al., the contents of which are incorporated herein by reference in their entirety.
  • the cationic lipid may have a positively-charged head group including two quaternary amine groups and a hydrophobic portion including four hydrocarbon chains as described in US Patent No. 7223887 to Gaucheron et al., the contents of which are incorporated herein by reference in their entirety.
  • the cationic lipid may be biodegradable as the biodegradable lipids disclosed in US20130195920 to Maier et al., the contents of which are incorporated herein by reference in their entirety.
  • the cationic lipid may have one or more biodegradable groups located in a lipidic moiety of the cationic lipid as described in formula I-IV in US 20130195920 to Maier et al., the contents of which are incorporated herein by reference in their entirety.
  • the cationic lipid may be selected from (20Z,23Z)- N,N-dimethylnonacosa-20,23-dien-10-amine, (17Z,20Z)-N,N-dimemylhexacosa-17,20-dien- 9-amine, (1Z,19Z)-N5N-dimethylpentacosa-l 6, 19-dien-8-amine, (13Z,16Z)-N,N- dimethyldocosa-13,16-dien-5-amine, (12Z,15Z)-N,N-dimethylhenicosa-12,15-dien-4-amine, (14Z,17Z)-N,N-dimethyltricosa-14,17-dien-6-amine, (15Z,18Z)-N,N-dimethyltetracosa- 15,18-dien-7-amine, (18Z,21Z)-N,N-dimethylheptacosa-18,21-dien-10-amine, (17Z,20Z)
  • the lipid may be a cleavable lipid such as those described in International Publication No. WO2012170889, the contents of which are herein incorporated by reference in their entirety.
  • the nanoparticles described herein may comprise at least one cationic polymer described herein and/or known in the art.
  • the cationic lipid may be synthesized by methods known in the art and/or as described in International Publication Nos.
  • the LNP formulations of the nucleic acid-based compositions may contain PEG-c-DOMG at 3% lipid molar ratio. In some embodiments, the LNP formulations of the nucleic acid-based compositions may contain PEG-c-DOMG at 1.5% lipid molar ratio.
  • the pharmaceutical compositions of the nucleic acid-based compositions may include at least one of the PEGylated lipids described in International Publication No. 2012099755, the contents of which is herein incorporated by reference in its entirety.
  • the LNP formulation may contain PEG-DMG 2000 (1,2- dimyristoyl-sn-glycero-3-phophoethanolamine-N-[methoxy(polyethylene glycol)-2000).
  • the LNP formulation may contain PEG-DMG 2000, a cationic lipid known in the art and at least one other component.
  • the LNP formulation may contain PEG-DMG 2000, a cationic lipid known in the art, DSPC and cholesterol.
  • the LNP formulation may contain PEG-DMG 2000, DLin-DMA, DSPC and cholesterol.
  • the LNP formulation may contain PEG-DMG 2000, DLin-DMA, DSPC and cholesterol in a molar ratio of 2:40:10:48 (see e.g., Geall et al., Non-viral delivery of self-amplifying RNA vaccines, PNAS 2012; PMID: 22908294; herein incorporated by reference in its entirety).
  • nucleic acid-based compositions described herein may be formulated in a nanoparticle to be delivered by a parenteral route as described in U.S. Pub. No. 20120207845; the contents of which is herein incorporated by reference in its entirety.
  • the cationic lipid may also be the cationic lipids disclosed in US20130156845 to Manoharan et al.
  • nucleic acid-based compositions of the present disclosure may be formulated with a plurality of cationic lipids, such as a first and a second cationic lipid as described in US20130017223 to Hope et al., the contents of which are incorporated herein by reference in their entirety.
  • the first cationic lipid can be selected on the basis of a first property and the second cationic lipid can be selected on the basis of a second property, where the properties may be determined as outlined in US20130017223, the contents of which are herein incorporated by reference in its entirety. In some embodiments, the first and second properties are complementary.
  • the nucleic acid-based compositions described herein may be formulated with a lipid particle comprising one or more cationic lipids and one or more second lipids, and one or more nucleic acids, wherein the lipid particle comprises a solid core, as described in US Patent Publication No. US20120276209 to Cullis et al., the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-based compositions of the present disclosure may be complexed with a cationic amphiphile in an oil-in-water (o/w) emulsion such as described in EP2298358 to Satishchandran et al., the contents of which are incorporated herein by reference in their entirety.
  • the cationic amphiphile may be a cationic lipid, modified or unmodified spermine, bupivacaine, or benzalkonium chloride and the oil may be a vegetable or an animal oil.
  • nucleic acid-based compositions of the present disclosure may be formulated with a composition comprising a mixture of cationic compounds and neutral lipids.
  • the cationic compounds may be formula (I) disclosed in WO 1999010390 to Ansell et al., the contents of which are disclosed herein by reference in their entirety
  • the neutral lipid may be selected from the group consisting of diacylphosphatidylcholine, diacylphosphatidylethanolamine, ceramide and sphingomyelin.
  • the lipid formulation may comprise a cationic lipid of formula A, a neutral lipid, a sterol and a PEG or PEG-modified lipid disclosed in US 20120101148 to Akinc et al., the contents of which are incorporated herein by reference in their entirety.
  • the LNP formulation may be formulated by the methods described in International Publication Nos. WO2011127255 or WO2008103276, each of which are herein incorporated by reference in their entirety.
  • the nucleic acid-based compositions of the present disclosure may be encapsulated in any of the lipid nanoparticle (LNP) formulations described in WO2011127255 and/or WO2008103276; the contents of each of which are herein incorporated by reference in their entirety.
  • the LNP formulations described herein may comprise a polycationic composition.
  • the polycationic composition may be selected from formula 1-60 of US Patent Publication No.
  • the LNP formulations comprising a polycationic composition may be used for the delivery of the nucleic acid-based compositions described herein in vivo and/or in vitro.
  • the LNP formulations described herein may additionally comprise a permeability enhancer molecule.
  • Non-limiting permeability enhancer molecules are described in US Patent Publication No. US20050222064; the contents of which is herein incorporated by reference in its entirety.
  • the pharmaceutical compositions may be formulated in liposomes such as, but not limited to, DiLa2 liposomes (Marina Biotech, Bothell, WA), SMARTICLES®/NOV340 (Marina Biotech, Bothell, WA), neutral DOPC (1,2-dioleoyl-sn- glycero-3-phosphocholine) based liposomes (e.g., siRNA delivery for ovarian cancer (Landen et al. Cancer Biology & Therapy 20065(12)1708-1713); the contents of which is herein incorporated by reference in its entirety) and hyaluronan-coated liposomes (Quiet Therapeutics, Israel).
  • DiLa2 liposomes Marina Biotech, Bothell, WA
  • SMARTICLES®/NOV340 Marina Biotech, Bothell, WA
  • neutral DOPC 1,2-dioleoyl-sn- glycero-3-phosphocholine
  • siRNA delivery for ovarian cancer Lianden
  • the pharmaceutical compositions may be formulated with any amphoteric liposome disclosed in WO 2008/043575 to Panzner and US 8580297 to Essler et al. (Marina Biotech), the contents of which are incorporated herein by reference in their entirety.
  • the amphoteric liposome may comprise a mixture of lipids including a cationic amphiphile, an anionic amphiphile and optional one or more neutral amphiphiles.
  • the amphoteric liposome may comprise amphoteric compounds based on amphiphilic molecules, the head groups of which being substituted with one or more amphoteric groups.
  • the pharmaceutical compositions may be formulated with an amphoteric lipid comprising one or more amphoteric groups having an isoelectric point between 4 and 9, as disclosed in US 20140227345 to Essler et al. (Marina Biotech), the contents of which are incorporated herein by reference in their entirety.
  • the pharmaceutical composition may be formulated with liposomes comprising a sterol derivative as disclosed in US 7312206 to Panzner et al. (Novosom), the contents of which are incorporated herein by reference in their entirety.
  • the pharmaceutical composition may be formulated with amphoteric liposomes comprising at least one amphipathic cationic lipid, at least one amphipathic anionic lipid, and at least one neutral lipid, or liposomes comprise at least one amphipathic lipid with both a positive and a negative charge, and at least one neutral lipid, wherein the liposomes are stable at pH 4.2 and pH 7.5, as disclosed in US Pat. No. 7780983 to Panzner et al. (Novosom), the contents of which are incorporated herein by reference in their entirety.
  • the pharmaceutical composition may be formulated with liposomes comprising a serum-stable mixture of lipids taught in US 20110076322 to Panzner et al, the contents of which are incorporated herein by reference in their entirety, capable of encapsulating the nucleic acid-based compositions of the present disclosure.
  • the lipid mixture comprises phosphatidylcholine and phosphatidylethanolamine in a ratio in the range of about 0.5 to about 8.
  • the lipid mixture may also include pH sensitive anionic and cationic amphiphiles, such that the mixture is amphoteric, being negatively charged or neutral at pH 7.4 and positively charged at pH 4.
  • the drug/lipid ratio may be adjusted to target the liposomes to particular organs or other sites in the body.
  • liposomes loaded with the nucleic acid-based compositions of the present disclosure as cargo are prepared by the method disclosed in US 20120021042 to Panzner et al., the contents of which are incorporated herein by reference in their entirety.
  • the method comprises steps of admixing an aqueous solution of a polyanionic active agent and an alcoholic solution of one or more amphiphiles and buffering said admixture to an acidic pH, wherein the one or more amphiphiles are susceptible of forming amphoteric liposomes at the acidic pH, thereby to form amphoteric liposomes in suspension encapsulating the active agent.
  • the nanoparticle formulations may be a carbohydrate nanoparticle comprising a carbohydrate carrier and a nucleic acid-based composition (e.g., an ASO).
  • the carbohydrate carrier may include, but is not limited to, an anhydride-modified phytoglycogen or glycogen-type material, phtoglycogen octenyl succinate, phytoglycogen beta-dextrin, anhydride-modified phytoglycogen beta-dextrin. (See e.g., International Publication No. WO2012109121; the contents of which is herein incorporated by reference in its entirety).
  • Lipid nanoparticle formulations may be improved by replacing the cationic lipid with a biodegradable cationic lipid which is known as a rapidly eliminated lipid nanoparticle (reLNP).
  • Ionizable cationic lipids such as, but not limited to, DLinDMA, DLin-KC2-DMA, and DLin-MC3-DMA, have been shown to accumulate in plasma and tissues over time and may be a potential source of toxicity.
  • the rapid metabolism of the rapidly eliminated lipids can improve the tolerability and therapeutic index of the lipid nanoparticles by an order of magnitude from a 1 mg/kg dose to a 10 mg/kg dose in rat.
  • ester linkage can improve the degradation and metabolism profile of the cationic component, while still maintaining the activity of the reLNP formulation.
  • the ester linkage can be internally located within the lipid chain or it may be terminally located at the terminal end of the lipid chain.
  • the internal ester linkage may replace any carbon in the lipid chain.
  • the nucleic acid-based compositions may be formulated as a lipoplex, such as, without limitation, the ATUPLEX TM system, the DACC system, the DBTC system and other siRNA-lipoplex technology from Silence Therapeutics (London, United Kingdom), STEMFECT TM from STEMGENT® (Cambridge, MA), and polyethylenimine (PEI) or protamine-based targeted and non-targeted delivery of nucleic acids (Aleku et al. Cancer Res. 200868:9788-9798; Strumberg et al.
  • a lipoplex such as, without limitation, the ATUPLEX TM system, the DACC system, the DBTC system and other siRNA-lipoplex technology from Silence Therapeutics (London, United Kingdom), STEMFECT TM from STEMGENT® (Cambridge, MA), and polyethylenimine (PEI) or protamine-based targeted and non-targeted delivery of nucleic acids (Aleku et al. Cancer Res. 200868
  • such formulations may also be constructed or compositions altered such that they passively or actively are directed to different cell types in vivo, including but not limited to hepatocytes, immune cells, tumor cells, endothelial cells, antigen presenting cells, and leukocytes (Akinc et al. Mol Ther. 201018:1357-1364; Song et al., Nat Biotechnol.
  • One example of passive targeting of formulations to liver cells includes the DLin- DMA, DLin-KC2-DMA and DLin-MC3-DMA-based lipid nanoparticle formulations which have been shown to bind to apolipoprotein E and promote binding and uptake of these formulations into hepatocytes in vivo (Akinc et al. Mol Ther. 201018:1357-1364; the contents of which is herein incorporated by reference in its entirety).
  • Formulations can also be selectively targeted through expression of different ligands on their surface as exemplified by, but not limited by, folate, transferrin, N-acetylgalactosamine (GalNAc), and antibody targeted approaches (Kolhatkar et al., Curr Drug Discov Technol. 20118:197-206; Musacchio and Torchilin, Front Biosci. 201116:1388-1412; Yu et al., Mol Membr Biol. 201027:286-298; Patil et al., Crit Rev Ther Drug Carrier Syst. 200825:1-61; Benoit et al., Biomacromolecules.
  • the nucleic acid-based compositions is formulated as a solid lipid nanoparticle.
  • a solid lipid nanoparticle may be spherical with an average diameter between 10 to 1000 nm.
  • SLN possess a solid lipid core matrix that can solubilize lipophilic molecules and may be stabilized with surfactants and/or emulsifiers.
  • the lipid nanoparticle may be a self-assembly lipid-polymer nanoparticle (see Zhang et al., ACS Nano, 2008, 2 (8), pp 1696–1702; the contents of which are herein incorporated by reference in its entirety).
  • controlled release refers to a pharmaceutical composition or compound release profile that conforms to a particular pattern of release to affect a therapeutic outcome.
  • the nucleic acid-based compositions may be encapsulated into a delivery agent described herein and/or known in the art for controlled release and/or targeted delivery.
  • the term “encapsulate” means to enclose, surround or encase. As it relates to the formulation of the compounds of the disclosure, encapsulation may be substantial, complete or partial.
  • the term “substantially encapsulated” means that at least greater than 50, 60, 70, 80, 85, 90, 95, 96, 97, 98, 99, 99.9, 99.9 or greater than 99.999% of the pharmaceutical composition or compound of the disclosure may be enclosed, surrounded or encased within the delivery agent.
  • “Partially encapsulated” means that less than 10, 10, 20, 30, 4050 or less of the pharmaceutical composition or compound of the disclosure may be enclosed, surrounded or encased within the delivery agent.
  • encapsulation may be determined by measuring the escape or the activity of the pharmaceutical composition or compound of the disclosure using fluorescence and/or electron micrograph. For example, at least 1, 5, 10, 20, 30, 40, 50, 60, 70, 80, 85, 90, 95, 96, 97, 98, 99, 99.9, 99.99 or greater than 99.99% of the pharmaceutical composition or compound of the disclosure are encapsulated in the delivery agent.
  • the nucleic acid-based compositions may be encapsulated into a lipid nanoparticle or a rapidly eliminated lipid nanoparticle and the lipid nanoparticles or a rapidly eliminated lipid nanoparticle may then be encapsulated into a polymer, hydrogel and/or surgical sealant described herein and/or known in the art.
  • the polymer, hydrogel or surgical sealant may be PLGA, ethylene vinyl acetate (EVAc), poloxamer, GELSITE® (Nanotherapeutics, Inc. Alachua, FL), HYLENEX® (Halozyme Therapeutics, San Diego CA), surgical sealants such as fibrinogen polymers (Ethicon Inc. Cornelia, GA), TISSELL® (Baxter International, Inc., Deerfield, IL), PEG-based sealants, and COSEAL® (Baxter International, Inc., Deerfield, IL).
  • EVAc ethylene vinyl acetate
  • poloxamer GELSITE®
  • GELSITE® Nanotherapeutics, Inc. Alachua, FL
  • HYLENEX® HyLENEX®
  • surgical sealants such as fibrinogen polymers (Ethicon Inc. Cornelia, GA), TISSELL® (Baxter International, Inc., Deerfield, IL), PEG-based sealants, and COSEAL
  • the lipid nanoparticle may be encapsulated into any polymer known in the art which may form a gel when injected into a subject.
  • the lipid nanoparticle may be encapsulated into a polymer matrix which may be biodegradable.
  • the nucleic acid-based compositions formulation for controlled release and/or targeted delivery may also include at least one controlled release coating.
  • Controlled release coatings include, but are not limited to, OPADRY®, polyvinylpyrrolidone/vinyl acetate copolymer, polyvinylpyrrolidone, hydroxypropyl methylcellulose, hydroxypropyl cellulose, hydroxyethyl cellulose, EUDRAGIT RL®, EUDRAGIT RS® and cellulose derivatives such as ethylcellulose aqueous dispersions (AQUACOAT® and SURELEASE®).
  • the controlled release and/or targeted delivery formulation may comprise at least one degradable polyester which may contain polycationic side chains.
  • Degradable polyesters include, but are not limited to, poly(serine ester), poly(L-lactide-co-L- lysine), poly(4-hydroxy-L-proline ester), and combinations thereof.
  • the degradable polyesters may include a PEG conjugation to form a PEGylated polymer.
  • the nucleic acid-based compositions of the present disclosure may be formulated with a targeting lipid with a targeting moiety such as the targeting moieties disclosed in US20130202652 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety.
  • the targeting moiety of formula I of US 20130202652 to Manoharan et al. may selected in order to favor the lipid being localized with a desired organ, tissue, cell, cell type or subtype, or organelle.
  • Non-limiting targeting moieties that are contemplated in the present disclosure include transferrin, anisamide, an RGD peptide, prostate specific membrane antigen (PSMA), fucose, an antibody, or an aptamer.
  • the nucleic acid-based compositions of the present disclosure may be encapsulated in a therapeutic nanoparticle. Therapeutic nanoparticles may be formulated by methods described herein and known in the art such as, but not limited to, International Pub Nos.
  • WO2010005740 WO2010030763, WO2010005721, WO2010005723, WO2012054923, US Pub. Nos. US20110262491, US20100104645, US20100087337, US20100068285, US20110274759, US20100068286 and US20120288541 and US Pat No. 8,206,747, 8,293,276, 8,318,208 and 8,318,211; the contents of each of which are herein incorporated by reference in their entirety.
  • Therapeutic polymer nanoparticles may be identified by the methods described in US Pub No. US20120140790, the contents of which are herein incorporated by reference in its entirety. [0219] In some embodiments, the therapeutic nanoparticle may be formulated for sustained release.
  • sustained release refers to a pharmaceutical composition or compound that conforms to a release rate over a specific period of time.
  • the period of time may include, but is not limited to, hours, days, weeks, months and years.
  • the sustained release nanoparticle may comprise a polymer and a therapeutic agent such as, but not limited to, the nucleic acid-based compositions of the present disclosure (see International Pub No. 2010075072 and US Pub No. US20100216804, US20110217377 and US20120201859, the contents of each of which are herein incorporated by reference in their entirety).
  • the therapeutic nanoparticles may be formulated to be target specific.
  • the therapeutic nanoparticles may include a corticosteroid (see International Pub. No. WO2011084518; the contents of which are herein incorporated by reference in its entirety).
  • the therapeutic nanoparticles may be formulated to be cancer specific.
  • the therapeutic nanoparticles may be formulated in nanoparticles described in International Pub No. WO2008121949, WO2010005726, WO2010005725, WO2011084521 and US Pub No. US20100069426, US20120004293 and US20100104655, the contents of each of which are herein incorporated by reference in their entirety.
  • the nanoparticles of the present disclosure may comprise a polymeric matrix.
  • the nanoparticle may comprise two or more polymers such as, but not limited to, polyethylenes, polycarbonates, polyanhydrides, polyhydroxyacids, polypropylfumerates, polycaprolactones, polyamides, polyacetals, polyethers, polyesters, poly(orthoesters), polycyanoacrylates, polyvinyl alcohols, polyurethanes, polyphosphazenes, polyacrylates, polymethacrylates, polycyanoacrylates, polyureas, polystyrenes, polyamines, polylysine, poly(ethylene imine), poly(serine ester), poly(L-lactide-co-L-lysine), poly(4-hydroxy-L-proline ester) or combinations thereof.
  • the therapeutic nanoparticle comprises a diblock copolymer.
  • the diblock copolymer may include PEG in combination with a polymer such as, but not limited to, polyethylenes, polycarbonates, polyanhydrides, polyhydroxyacids, polypropylfumerates, polycaprolactones, polyamides, polyacetals, polyethers, polyesters, poly(orthoesters), polycyanoacrylates, polyvinyl alcohols, polyurethanes, polyphosphazenes, polyacrylates, polymethacrylates, polycyanoacrylates, polyureas, polystyrenes, polyamines, polylysine, poly(ethylene imine), poly(serine ester), poly(L-lactide-co-L-lysine), poly(4-hydroxy-L-proline ester) or combinations thereof.
  • a polymer such as, but not limited to, polyethylenes, polycarbonates, polyanhydrides, polyhydroxyacids, polypropylfumerates
  • the therapeutic nanoparticle comprises a PLGA-PEG block copolymer (see US Pub. No. US20120004293 and US Pat No. 8,236,330, each of which is herein incorporated by reference in their entirety).
  • the therapeutic nanoparticle is a stealth nanoparticle comprising a diblock copolymer of PEG and PLA or PEG and PLGA (see US Pat No 8,246,968 and International Publication No. WO2012166923, the contents of each of which is herein incorporated by reference in its entirety).
  • the therapeutic nanoparticle may comprise a multiblock copolymer such as, but not limited to the multiblock copolymers described in U.S.
  • the block copolymers described herein may be included in a polyion complex comprising a non-polymeric micelle and the block copolymer. (See e.g., U.S. Pub. No. 20120076836; the contents of which are herein incorporated by reference in its entirety).
  • the therapeutic nanoparticle may comprise at least one acrylic polymer.
  • Acrylic polymers include but are not limited to, acrylic acid, methacrylic acid, acrylic acid and methacrylic acid copolymers, methyl methacrylate copolymers, ethoxyethyl methacrylates, cyanoethyl methacrylate, amino alkyl methacrylate copolymer, poly(acrylic acid), poly(methacrylic acid), polycyanoacrylates and combinations thereof.
  • the therapeutic nanoparticles may comprise at least one amine-containing polymer such as, but not limited to polylysine, polyethylene imine, poly(amidoamine) dendrimers, poly(beta-amino esters) (See e.g., U.S. Pat. No.
  • the therapeutic nanoparticles may comprise at least one degradable polyester which may contain polycationic side chains.
  • Degradable polyesters include, but are not limited to, poly(serine ester), poly(L-lactide-co-L-lysine), poly(4- hydroxy-L-proline ester), and combinations thereof.
  • the degradable polyesters may include a PEG conjugation to form a PEGylated polymer.
  • the therapeutic nanoparticle may include a conjugation of at least one targeting ligand.
  • the targeting ligand may be any ligand known in the art such as, but not limited to, a monoclonal antibody. (Kirpotin et al, Cancer Res. 200666:6732-6740; the contents of which are herein incorporated by reference in its entirety).
  • the therapeutic nanoparticle may be formulated in an aqueous solution which may be used to target cancer (see International Pub No. WO2011084513 and US Pub No. US20110294717, the contents of each of which is herein incorporated by reference in their entirety).
  • the nucleic acid-based compositions may be encapsulated in, linked to and/or associated with synthetic nanocarriers.
  • Synthetic nanocarriers include, but are not limited to, those described in International Pub. Nos. WO2010005740, WO2010030763, WO201213501, WO2012149252, WO2012149255, WO2012149259, WO2012149265, WO2012149268, WO2012149282, WO2012149301, WO2012149393, WO2012149405, WO2012149411, WO2012149454 and WO2013019669, and US Pub. Nos. US20110262491, US20100104645, US20100087337 and US20120244222, the contents of each of which are herein incorporated by reference in their entirety.
  • the synthetic nanocarriers may be formulated using methods known in the art and/or described herein.
  • the synthetic nanocarriers may be formulated by the methods described in International Pub Nos. WO2010005740, WO2010030763 and WO201213501and US Pub. Nos. US20110262491, US20100104645, US20100087337 and US2012024422, the contents of each of which are herein incorporated by reference in their entirety.
  • the synthetic nanocarrier formulations may be lyophilized by methods described in International Pub. No. WO2011072218 and US Pat No. 8,211,473; the contents of each of which are herein incorporated by reference in their entirety.
  • the synthetic nanocarriers may contain reactive groups to release the nucleic acid-based compositions described herein (see International Pub.
  • the synthetic nanocarriers may be formulated for targeted release.
  • the synthetic nanocarrier may be formulated to release the nucleic acid-based compositions at a specified pH and/or after a desired time interval.
  • the synthetic nanoparticle may be formulated to release the nucleic acid-based compositions after 24 hours and/or at a pH of 4.5 (see International Pub. Nos. WO2010138193 and WO2010138194 and US Pub Nos.
  • the synthetic nanocarriers may be formulated for controlled and/or sustained release of the nucleic acid-based compositions described herein.
  • the synthetic nanocarriers for sustained release may be formulated by methods known in the art, described herein and/or as described in International Pub No. WO2010138192 and US Pub No. 20100303850, the contents each of which is herein incorporated by reference in their entirety.
  • the nanoparticle may be optimized for oral administration.
  • the nanoparticle may comprise at least one cationic biopolymer such as, but not limited to, chitosan or a derivative thereof.
  • the nanoparticle may be formulated by the methods described in U.S. Pub. No. 20120282343; the contents of which are herein incorporated by reference in its entirety.
  • the nucleic acid-based compositions of the present disclosure may be formulated in a modular composition such as described in US 8575123 to Manoharan et al., the contents of which are herein incorporated by reference in their entirety.
  • the modular composition may comprise a nucleic acid, e.g., the nucleic acid-based compositions of the present disclosure, at least one endosomolytic component, and at least one targeting ligand.
  • the modular composition may have a formula such as any formula described in US 8575123 to Manoharan et al., the contents of which are herein incorporated by reference in their entirety.
  • the nucleic acid-based compositions of the present disclosure may be encapsulated in the lipid formulation to form a stable nucleic acid-lipid particle (SNALP) such as described in US8546554 to de Fougerolles et al., the contents of which are incorporated here by reference in their entirety.
  • SNALP stable nucleic acid-lipid particle
  • the lipid may be cationic or non- cationic.
  • the lipid to nucleic acid ratio (mass/mass ratio) (e.g., lipid to nucleic acid-based compositions ratio) will be in the range of from about 1:1 to about 50:1, from about 1:1 to about 25:1, from about 3:1 to about 15:1, from about 4:1 to about 10:1, from about 5:1 to about 9:1, or about 6:1 to about 9:1, or 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, or 11:1.
  • the SNALP includes 40% 2,2-Dilinoleyl-4-dimethylaminoethyl- [1,3]-dioxolane (Lipid A), 10% dioleoylphosphatidylcholine (DSPC), 40% cholesterol, 10% polyethylene glycol (PEG)-C-DOMG (mole percent) with a particle size of 63.0 ⁇ 20 nm and a 0.027 nucleic acid/lipid ratio.
  • the nucleic acid-based compositions of the present disclosure may be formulated with a nucleic acid-lipid particle comprising an endosomal membrane destabilizer as disclosed in US 7189705 to Lam et al., the contents of which are incorporated herein by reference in their entirety.
  • the endosomal membrane destabilizer may be a Ca 2+ ion.
  • the nucleic acid-based compositions of the present disclosure may be formulated with formulated lipid particles (FLiPs) disclosed in US 8148344 to Akinc et al., the contents of which are herein incorporated by reference in their entirety. Akinc et al.
  • FLiPs may comprise at least one of a single or double- stranded oligonucleotide, where the oligonucleotide has been conjugated to a lipophile and at least one of an emulsion or liposome to which the conjugated oligonucleotide has been aggregated, admixed or associated.
  • These particles have surprisingly been shown to effectively deliver oligonucleotides to heart, lung and muscle disclosed in US 8148344 to Akinc et al., the contents of which are herein incorporated by reference in their entirety.
  • the nucleic acid-based compositions of the present disclosure may be delivered to a cell using a composition comprising an expression vector in a lipid formulation as described in US 6086913 to Tam et al., the contents of which are incorporated herein by reference in their entirety.
  • the composition disclosed by Tam is serum-stable and comprises an expression vector comprising first and second inverted repeated sequences from an adeno associated virus (AAV), a rep gene from AAV, and a nucleic acid fragment.
  • AAV adeno associated virus
  • the expression vector in Tam is complexed with lipids.
  • the nucleic acid-based compositions of the present disclosure may be formulated with a lipid formulation disclosed in US 20120270921 to de Fougerolles et al., the contents of which are incorporated herein by reference in their entirety.
  • the lipid formulation may include a cationic lipid having the formula A described in US 20120270921, the contents of which are herein incorporated by reference in its entirety.
  • the compositions of exemplary nucleic acid-lipid particles disclosed in Table A of US20120270921, the contents of which are incorporated herein by reference in their entirety may be used with the nucleic acid-based compositions of the present disclosure.
  • the nucleic acid-based compositions of the present disclosure may be fully encapsulated in a lipid particle disclosed in US 20120276207 to Maurer et al., the contents of which are incorporated herein by reference in their entirety.
  • the particles may comprise a lipid composition comprising preformed lipid vesicles, a charged therapeutic agent, and a destabilizing agent to form a mixture of preformed vesicles and therapeutic agent in a destabilizing solvent, wherein the destabilizing solvent is effective to destabilize the membrane of the preformed lipid vesicles without disrupting the vesicles.
  • the nucleic acid-based compositions of the present disclosure may be formulated with a conjugated lipid.
  • the conjugated lipid may have a formula such as described in US 20120264810 to Lin et al., the contents of which are incorporated herein by reference in their entirety.
  • the conjugate lipid may form a lipid particle which further comprises a cationic lipid, a neutral lipid, and a lipid capable of reducing aggregation.
  • nucleic acid-based compositions of the present disclosure may be formulated in a neutral liposomal formulation such as disclosed in US 20120244207 to Fitzgerald et al., the contents of which are incorporated herein by reference in their entirety.
  • neutral liposomal formulation refers to a liposomal formulation with a near neutral or neutral surface charge at a physiological pH.
  • Physiological pH can be, e.g., about 7.0 to about 7.5, or, e.g., about 7.5, or, e.g., 7.0, 7.1, 7.2, 7.3, 7.4, or 7.5, or, e.g., 7.3, or, e.g., 7.4.
  • a neutral liposomal formulation is an ionizable lipid nanoparticle (iLNP).
  • a neutral liposomal formulation can include an ionizable cationic lipid, e.g., DLin-KC2-DMA.
  • the nucleic acid-based compositions of the present disclosure may be formulated with a charged lipid or an amino lipid.
  • charged lipid is meant to include those lipids having one or two fatty acyl or fatty alkyl chains and a quaternary amino head group. The quaternary amine carries a permanent positive charge.
  • the head group can optionally include an ionizable group, such as a primary, secondary, or tertiary amine that may be protonated at physiological pH.
  • an ionizable group such as a primary, secondary, or tertiary amine that may be protonated at physiological pH.
  • the presence of the quaternary amine can alter the pKa of the ionizable group relative to the pKa of the group in a structurally similar compound that lacks the quaternary amine (e.g., the quaternary amine is replaced by a tertiary amine)
  • a charged lipid is referred to as an "amino lipid.”
  • the amino lipid may be any amino lipid described in US20110256175 to Hope et al., the contents of which are incorporated herein by reference in their entirety.
  • the amino lipids may have the structure disclosed in Tables 3-7 of Hope, such as structure (II), DLin-K-C2-DMA, DLin-K2-DMA, DLin-K6- DMA, etc.
  • the resulting pharmaceutical preparations may be lyophilized according to Hope.
  • the amino lipids may be any amino lipid described in US 20110117125 to Hope et al., the contents of which are incorporated herein by reference in their entirety, such as a lipid of structure (I), DLin-K-DMA, DLin-C-DAP, DLin-DAC, DLin-MA, DLin-S-DMA, etc.
  • the amino lipid may have the structure (I), (II), (III), or (IV), or 4-(R)-DUn-K-DMA (VI), 4-(S)-DUn-K-DMA (V) as described in WO2009132131 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety.
  • the charged lipid used in any of the formulations described herein may be any charged lipid described in EP2509636 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-based composition s of the present disclosure may be formulated with an association complex containing lipids, liposomes, or lipoplexes.
  • the association complex comprises one or more compounds each having a structure defined by formula (I), a PEG-lipid having a structure defined by formula (XV), a steroid and a nucleic acid disclosed in US8034376 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-based compositions may be formulated with any association complex described in US8034376, the contents of which are herein incorporated by reference in its entirety.
  • the nucleic acid-based compositions of the present disclosure may be formulated with reverse head group lipids.
  • the nucleic acid-based compositions may be formulated with a zwitterionic lipid comprising a headgroup wherein the positive charge is located near the acyl chain region and the negative charge is located at the distal end of the head group, such as a lipid having structure (A) or structure (I) described in WO2011056682 to Leung et al., the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-based compositions of the present disclosure may be formulated in a lipid bilayer carrier.
  • the nucleic acid-based compositions may be combined with a lipid-detergent mixture comprising a lipid mixture of an aggregation-preventing agent in an amount of about 5 mol% to about 20 mol%, a cationic lipid in an amount of about 0.5 mol% to about 50 mol%, and a fusogenic lipid and a detergent, to provide a nucleic acid-lipid-detergent mixture; and then dialyzing the nucleic acid-lipid-detergent mixture against a buffered salt solution to remove the detergent and to encapsulate the nucleic acid in a lipid bilayer carrier and provide a lipid bilayer-nucleic acid composition, wherein the buffered salt solution has an ionic strength sufficient to encapsulate of from about 40 % to about 80 % of the nucleic acid, described in WO1999018933 to Cullis et al., the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-based compositions of the present disclosure may be formulated in a nucleic acid-lipid particle capable of selectively targeting the nucleic acid-based compositions to a kidney, heart, liver, or tumor tissue site.
  • the nucleic acid-lipid particle may comprise (a) a nucleic acid; (b) 1.0 mole % to 45 mole % of a cationic lipid; (c) 0,0 mole % to 90 mole % of another lipid; (d) 1,0 mole % to 10 mole % of a bilayer stabilizing component; (e) 0,0 mole % to 60 mole % cholesterol; and (f) 0,0 mole % to 10 mole % of cationic polymer lipid as described in EP1328254 to Cullis et al., the contents of which are incorporated herein by reference in their entirety.
  • Cullis teaches that varying the amount of each of the cationic lipid, bilayer stabilizing component, another lipid, cholesterol, and cationic polymer lipid can impart tissue selectivity for heart, liver, or tumor tissue site, thereby identifying a nucleic acid-lipid particle capable of selectively targeting a nucleic acid to the heart, liver, or tumor tissue site.
  • Polymers, Biodegradable Nanoparticles, and Core-Shell Nanoparticles [0249]
  • the nucleic acid-based compositions of the disclosure can be formulated using natural and/or synthetic polymers.
  • Non-limiting examples of polymers which may be used for delivery include, but are not limited to, DYNAMIC POLYCONJUGATE® (Arrowhead Research Corp., Pasadena, CA) formulations from MIRUS® Bio (Madison, WI) and Roche Madison (Madison, WI), PHASERX TM polymer formulations such as, without limitation, SMARTT POLYMER TECHNOLOGYTM (PHASERX®, Seattle, WA), DMRI/DOPE, poloxamer, VAXFECTIN® adjuvant from Vical (San Diego, CA), chitosan, cyclodextrin from Calando Pharmaceuticals (Pasadena, CA), dendrimers and poly(lactic-co-glycolic acid) (PLGA) polymers.
  • DYNAMIC POLYCONJUGATE® Arrowhead Research Corp., Pasadena, CA
  • PHASERX TM polymer formulations such as, without limitation, SMARTT POLYMER TECHNOLOGYTM (
  • chitosan formulation includes a core of positively charged chitosan and an outer portion of negatively charged substrate (U.S. Pub. No. 20120258176; herein incorporated by reference in its entirety).
  • Chitosan includes, but is not limited to N-trimethyl chitosan, mono-N-carboxymethyl chitosan (MCC), N-palmitoyl chitosan (NPCS), EDTA-chitosan, low molecular weight chitosan, chitosan derivatives, or combinations thereof.
  • MCC mono-N-carboxymethyl chitosan
  • NPCS N-palmitoyl chitosan
  • EDTA-chitosan low molecular weight chitosan
  • chitosan derivatives or combinations thereof.
  • the polymers used in the present disclosure have undergone processing to reduce and/or inhibit the attachment of unwanted substances such as, but not limited to, bacteria, to the surface of the polymer.
  • the polymer may be processed by methods known and/or described in the art and/or described in International Pub. No. WO2012150467, herein incorporated by reference in its entirety.
  • a non-limiting example of PLGA formulations include, but are not limited to, PLGA injectable depots (e.g., ELIGARD® which is formed by dissolving PLGA in 66% N- methyl-2-pyrrolidone (NMP) and the remainder being aqueous solvent and leuprolide. Once injected, the PLGA and leuprolide peptide precipitates into the subcutaneous space).
  • PLGA injectable depots e.g., ELIGARD® which is formed by dissolving PLGA in 66% N- methyl-2-pyrrolidone (NMP) and the remainder being aqueous solvent and leuprolide. Once injected, the PLGA and leuprolide peptide precipitates into the subcutaneous space).
  • NMP N- methyl-2-pyrrolidone
  • siRNA small interfering RNA
  • RNA small interfering RNA
  • the first of these delivery approaches uses dynamic polyconjugates and has been shown in vivo in mice to effectively deliver siRNA and silence endogenous target mRNA in hepatocytes (Rozema et al., Proc Natl Acad Sci U S A. 2007104:12982-12887; herein incorporated by reference in its entirety).
  • This particular approach is a multicomponent polymer system whose key features include a membrane-active polymer to which nucleic acid, in this case siRNA, is covalently coupled via a disulfide bond and where both PEG (for charge masking) and N-acetylgalactosamine (for hepatocyte targeting) groups are linked via pH-sensitive bonds (Rozema et al., Proc Natl Acad Sci U S A. 2007104:12982-12887; herein incorporated by reference in its entirety).
  • the polymer complex On binding to the hepatocyte and entry into the endosome, the polymer complex disassembles in the low-pH environment, with the polymer exposing its positive charge, leading to endosomal escape and cytoplasmic release of the siRNA from the polymer.
  • the polymer Through replacement of the N-acetylgalactosamine group with a mannose group, it was shown one could alter targeting from asialoglycoprotein receptor-expressing hepatocytes to sinusoidal endothelium and Kupffer cells.
  • Another polymer approach involves using transferrin-targeted cyclodextrin-containing polycation nanoparticles.
  • the polymer formulation can permit the sustained or delayed release of nucleic acid-based compositions (e.g., following intramuscular, subcutaneous, intraparenchymal, intrathecal, intracerebroventricular administration).
  • the altered release profile for the nucleic acid-based compositions can result in, for example, translation of an encoded protein over an extended period of time.
  • Biodegradable polymers have been previously used to protect nucleic acids from degradation and been shown to result in sustained release of payloads in vivo (Rozema et al., Proc Natl Acad Sci U S A. 2007104:12982-12887; Sullivan et al., Expert Opin Drug Deliv.
  • the pharmaceutical compositions may be sustained release formulations.
  • the sustained release formulations may be for subcutaneous delivery.
  • Sustained release formulations may include, but are not limited to, PLGA microspheres, ethylene vinyl acetate (EVAc), poloxamer, GELSITE® (Nanotherapeutics, Inc. Alachua, FL), HYLENEX® (Halozyme Therapeutics, San Diego CA), surgical sealants such as fibrinogen polymers (Ethicon Inc.
  • nucleic acid-based compositions may be formulated in PLGA microspheres by preparing the PLGA microspheres with tunable release rates (e.g., days and weeks) and encapsulating the nucleic acid-based compositions in the PLGA microspheres while maintaining the integrity of the nucleic acid-based compositions during the encapsulation process.
  • tunable release rates e.g., days and weeks
  • EVAc are non-biodegradable, biocompatible polymers which are used extensively in pre-clinical sustained release implant applications (e.g., extended release products Ocusert a pilocarpine ophthalmic insert for glaucoma or progestasert a sustained release progesterone intrauterine device; transdermal delivery systems Testoderm, Duragesic and Selegiline; catheters).
  • Poloxamer F-407 NF is a hydrophilic, non-ionic surfactant triblock copolymer of polyoxyethylene-polyoxypropylene-polyoxyethylene having a low viscosity at temperatures less than 5oC and forms a solid gel at temperatures greater than 15oC.
  • PEG- based surgical sealants comprise two synthetic PEG components mixed in a delivery device which can be prepared in one minute, seals in 3 minutes and is reabsorbed within 30 days.
  • GELSITE® and natural polymers are capable of in-situ gelation at the site of administration. They have been shown to interact with protein and peptide therapeutic candidates through ionic interaction to provide a stabilizing effect.
  • Polymer formulations can also be selectively targeted through expression of different ligands as exemplified by, but not limited by, folate, transferrin, and N- acetylgalactosamine (GalNAc) (Benoit et al., Biomacromolecules.
  • nucleic acid-based compositions of the disclosure may be formulated with or in a polymeric compound.
  • the polymer may include at least one polymer such as, but not limited to, polyethenes, polyethylene glycol (PEG), poly(l-lysine)(PLL), PEG grafted to PLL, cationic lipopolymer, biodegradable cationic lipopolymer, polyethyleneimine (PEI), cross- linked branched poly(alkylene imines), a polyamine derivative, a modified poloxamer, a biodegradable polymer, elastic biodegradable polymer, biodegradable block copolymer, biodegradable random copolymer, biodegradable polyester copolymer, biodegradable polyester block copolymer, biodegradable polyester block random copolymer, multiblock copolymers, linear biodegradable copolymer, poly[ ⁇ -(4-aminobutyl)-L-glycolic acid) (PAGA), biodegradable cross-linked cationic multi-block copolymers, polycarbonates, polyanhydrides, polyhydroxya
  • the nucleic acid-based compositions of the disclosure may be formulated with the polymeric compound of PEG grafted with PLL as described in U.S. Pat. No. 6,177,274; herein incorporated by reference in its entirety.
  • the formulation may be used for transfecting cells in vitro or for in vivo delivery of the nucleic acid-based compositions.
  • the nucleic acid-based compositions may be suspended in a solution or medium with a cationic polymer, in a dry pharmaceutical composition or in a solution that is capable of being dried as described in U.S. Pub. Nos. 20090042829 and 20090042825; each of which are herein incorporated by reference in their entireties.
  • nucleic acid-based compositions of the disclosure may be formulated with a PLGA-PEG block copolymer (see US Pub. No. US20120004293 and US Pat No. 8,236,330, herein incorporated by reference in their entireties) or PLGA-PEG-PLGA block copolymers (See U.S. Pat. No. 6,004,573, herein incorporated by reference in its entirety).
  • nucleic acid-based compositions of the disclosure may be formulated with a diblock copolymer of PEG and PLA or PEG and PLGA (see US Pat No 8,246,968, herein incorporated by reference in its entirety).
  • a polyamine derivative may be used to deliver nucleic acids or to treat and/or prevent a disease or to be included in an implantable or injectable device (U.S. Pub. No. 20100260817 herein incorporated by reference in its entirety).
  • a pharmaceutical composition may include the nucleic acid-based compositions and the polyamine derivative described in U.S. Pub. No. 20100260817, the contents of which are incorporated herein by reference in their entirety.
  • the nucleic acid-based compositions of the present disclosure may be delivered using a polyamide polymer such as, but not limited to, a polymer comprising a 1,3-dipolar addition polymer prepared by combining a carbohydrate diazide monomer with a dialkyne unite comprising oligoamines (U.S. Pat. No. 8,236,280; herein incorporated by reference in its entirety).
  • a polyamide polymer such as, but not limited to, a polymer comprising a 1,3-dipolar addition polymer prepared by combining a carbohydrate diazide monomer with a dialkyne unite comprising oligoamines (U.S. Pat. No. 8,236,280; herein incorporated by reference in its entirety).
  • the nucleic acid-based compositions of the present disclosure may be formulated with at least one polymer and/or derivatives thereof described in International Publication Nos. WO2011115862, WO2012082574 and WO2012068187 and U.S.
  • the nucleic acid-based compositions of the present disclosure may be formulated with a polymer of formula Z as described in WO2011115862, herein incorporated by reference in its entirety.
  • the nucleic acid-based compositions may be formulated with a polymer of formula Z, Z’ or Z’’ as described in International Pub. Nos. WO2012082574 or WO2012068187 and U.S. Pub. No. 2012028342, the contents of each of which are herein incorporated by reference in their entireties.
  • the polymers formulated with the nucleic acid-based compositions of the present disclosure may be synthesized by the methods described in International Pub. Nos.
  • the nucleic acid-based compositions of the disclosure may be formulated with at least one acrylic polymer.
  • Acrylic polymers include but are not limited to, acrylic acid, methacrylic acid, acrylic acid and methacrylic acid copolymers, methyl methacrylate copolymers, ethoxyethyl methacrylates, cyanoethyl methacrylate, amino alkyl methacrylate copolymer, poly(acrylic acid), poly(methacrylic acid), polycyanoacrylates and combinations thereof.
  • nucleic acid-based compositions of the disclosure may include at least one amine-containing polymer such as, but not limited to polylysine, polyethylene imine, poly(amidoamine) dendrimers or combinations thereof.
  • the nucleic acid-based compositions of the disclosure may be formulated in a pharmaceutical compound including a poly(alkylene imine), a biodegradable cationic lipopolymer, a biodegradable block copolymer, a biodegradable polymer, or a biodegradable random copolymer, a biodegradable polyester block copolymer, a biodegradable polyester polymer, a biodegradable polyester random copolymer, a linear biodegradable copolymer, PAGA, a biodegradable cross-linked cationic multi-block copolymer or combinations thereof.
  • the biodegradable cationic lipopolymer may be made by methods known in the art and/or described in U.S. Pat. No. 6,696,038, U.S. App. Nos. 20030073619 and 20040142474 each of which is herein incorporated by reference in their entireties.
  • the poly(alkylene imine) may be made using methods known in the art and/or as described in U.S. Pub. No. 20100004315, herein incorporated by reference in its entirety.
  • biodegradable polymer, biodegradable block copolymer, the biodegradable random copolymer, biodegradable polyester block copolymer, biodegradable polyester polymer, or biodegradable polyester random copolymer may be made using methods known in the art and/or as described in U.S. Pat. Nos. 6,517,869 and 6,267,987, the contents of which are each incorporated herein by reference in their entirety.
  • the linear biodegradable copolymer may be made using methods known in the art and/or as described in U.S. Pat. No. 6,652,886.
  • the PAGA polymer may be made using methods known in the art and/or as described in U.S. Pat. No.
  • the PAGA polymer may be copolymerized to form a copolymer or block copolymer with polymers such as but not limited to, poly-L-lysine, polyargine, polyornithine, histones, avidin, protamines, polylactides and poly(lactide-co-glycolides).
  • the biodegradable cross-linked cationic multi-block copolymers may be made my methods known in the art and/or as described in U.S. Pat. No. 8,057,821 or U.S. Pub. No. 2012009145 each of which are herein incorporated by reference in their entireties.
  • the multi-block copolymers may be synthesized using linear polyethyleneimine (LPEI) blocks which have distinct patterns as compared to branched polyethyleneimines.
  • LPEI linear polyethyleneimine
  • the composition or pharmaceutical composition may be made by the methods known in the art, described herein, or as described in U.S. Pub. No. 20100004315 or U.S. Pat. Nos. 6,267,987 and 6,217,912 each of which are herein incorporated by reference in their entireties.
  • the nucleic acid-based compositions of the disclosure may be formulated with at least one degradable polyester which may contain polycationic side chains.
  • Degradable polyesters include, but are not limited to, poly(serine ester), poly(L-lactide-co-L-lysine), poly(4-hydroxy-L-proline ester), and combinations thereof.
  • the degradable polyesters may include a PEG conjugation to form a PEGylated polymer.
  • the nucleic acid-based compositions of the disclosure may be formulated with at least one crosslinkable polyester.
  • Crosslinkable polyesters include those known in the art and described in US Pub. No. 20120269761, herein incorporated by reference in its entirety.
  • the polymers described herein may be conjugated to a lipid- terminating PEG.
  • PLGA may be conjugated to a lipid-terminating PEG forming PLGA-DSPE-PEG.
  • PEG conjugates for use with the present disclosure are described in International Publication No. WO2008103276, herein incorporated by reference in its entirety.
  • the polymers may be conjugated using a ligand conjugate such as, but not limited to, the conjugates described in U.S. Pat. No. 8,273,363, herein incorporated by reference in its entirety.
  • the nucleic acid-based compositions described herein may be conjugated with another compound.
  • Non-limiting examples of conjugates are described in US Patent Nos.
  • nucleic acid-based compositions of the present disclosure may be conjugated with conjugates of formula 1-122 as described in US Patent Nos. 7,964,578 and 7,833,992, each of which are herein incorporated by reference in their entireties.
  • the nucleic acid-based compositions described herein may be conjugated with a metal such as, but not limited to, gold. (See e.g., Giljohann et al. Journ. Amer. Chem. Soc. 2009131(6): 2072-2073; herein incorporated by reference in its entirety).
  • the nucleic acid-based compositions described herein may be conjugated and/or encapsulated in gold-nanoparticles.
  • a gene delivery composition may include a nucleotide sequence and a poloxamer.
  • the nucleic acid-based compositions of the present disclosure may be used in a gene delivery composition with the poloxamer described in U.S. Pub. No. 20100004313.
  • the polymer formulation of the present disclosure may be stabilized by contacting the polymer formulation, which may include a cationic carrier, with a cationic lipopolymer which may be covalently linked to cholesterol and polyethylene glycol groups.
  • the polymer formulation may be contacted with a cationic lipopolymer using the methods described in U.S. Pub. No. 20090042829 herein incorporated by reference in its entirety.
  • the cationic carrier may include, but is not limited to, polyethylenimine, poly(trimethylenimine), poly(tetramethylenimine), polypropylenimine, aminoglycoside- polyamine, dideoxy-diamino-b-cyclodextrin, spermine, spermidine, poly(2- dimethylamino)ethyl methacrylate, poly(lysine), poly(histidine), poly(arginine), cationized gelatin, dendrimers, chitosan, 1,2-Dioleoyl-3-Trimethylammonium-Propane(DOTAP), N-[1- (2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA), 1-[2- (oleoyloxy)ethyl]-2-oleyl-3-(2-hydroxyethyl)imidazolinium chloride (DOTIM), 2,3- dioleyloxy-
  • the nucleic acid-based compositions of the disclosure may be formulated in a polyplex of one or more polymers (U.S. Pub. No. 20120237565 and 20120270927; each of which is herein incorporated by reference in its entirety).
  • the polyplex comprises two or more cationic polymers.
  • the cationic polymer may comprise a poly(ethylene imine) (PEI) such as linear PEI.
  • PEI poly(ethylene imine)
  • the nucleic acid-based compositions of the disclosure can also be formulated as a nanoparticle using a combination of polymers, lipids, and/or other biodegradable agents, such as, but not limited to, calcium phosphate.
  • Components may be combined in a core-shell, hybrid, and/or layer-by-layer architecture, to allow for fine-tuning of the nanoparticle so delivery of the nucleic acid-based compositions may be enhanced (Wang et al., Nat Mater. 20065:791-796; Fuller et al., Biomaterials.200829:1526-1532; DeKoker et al., Adv Drug Deliv Rev. 201163:748-761; Endres et al., Biomaterials. 201132:7721-7731; Su et al., Mol Pharm. 2011 Jun 6;8(3):774-87; herein incorporated by reference in its entirety).
  • the nanoparticle may comprise a plurality of polymers such as, but not limited to hydrophilic-hydrophobic polymers (e.g., PEG-PLGA), hydrophobic polymers (e.g., PEG) and/or hydrophilic polymers (International Pub. No. WO20120225129; herein incorporated by reference in its entirety).
  • PEG-PLGA hydrophilic-hydrophobic polymers
  • PEG hydrophobic polymers
  • hydrophilic polymers International Pub. No. WO20120225129; herein incorporated by reference in its entirety.
  • Biodegradable calcium phosphate nanoparticles in combination with lipids and/or polymers may be used to deliver nucleic acid-based compositions in vivo.
  • a lipid coated calcium phosphate nanoparticle which may also contain a targeting ligand such as anisamide, may be used to deliver the nucleic acid-based compositions of the present disclosure.
  • a targeting ligand such as anisamide
  • a lipid coated calcium phosphate nanoparticle was used (Li et al., J Contr Rel. 2010142: 416-421; Li et al., J Contr Rel. 2012158:108-114; Yang et al., Mol Ther. 201220:609-615; herein incorporated by reference in its entirety).
  • This delivery system combines both a targeted nanoparticle and a component to enhance the endosomal escape, calcium phosphate, in order to improve delivery of the siRNA.
  • calcium phosphate with a PEG-polyanion block copolymer may be used to delivery nucleic acid-based compositions (Kazikawa et al., J Contr Rel. 2004 97:345-356; Kazikawa et al., J Contr Rel. 2006111:368-370; herein incorporated by reference in its entirety).
  • a PEG-charge-conversional polymer (Pitella et al., Biomaterials.
  • the PEG-charge-conversional polymer may improve upon the PEG-polyanion block copolymers by being cleaved into a polycation at acidic pH, thus enhancing endosomal escape.
  • the use of core-shell nanoparticles has additionally focused on a high-throughput approach to synthesize cationic cross-linked nanogel cores and various shells (Siegwart et al., Proc Natl Acad Sci U S A. 2011108:12996-13001).
  • the complexation, delivery, and internalization of the polymeric nanoparticles can be precisely controlled by altering the chemical composition in both the core and shell components of the nanoparticle.
  • the core-shell nanoparticles may efficiently deliver nucleic acid-based compositions to mouse hepatocytes after they covalently attach cholesterol to the nanoparticle.
  • the nanoparticles described herein may be nanoparticles which include at least one ligand, and the ligand may be a peptide which targets tumors (i.e., a targeting peptide).
  • the targeting peptide is a cyclic RGD (cRGD) ligand which was found to bind to ⁇ v ⁇ 3 and ⁇ v ⁇ 5 integrins which are highly expressed on angiogenic endothelial cells of tumors as well as on a number of tumor cells.
  • the targeting peptide is an APRPG peptide (SEQ ID NO: 24457) which is an angiogenic vessel-homing peptide.
  • the targeting peptide is a cyclic NGR (CNGRCVSGCAGRC provided as SEQ ID NO: 24445) which is upregulated primarily in the tumor vasculature and in a few cancers such as fibrosarcomas.
  • Linear NGR may also be an option for a targeting peptide.
  • the targeting peptide is a F3 peptide (KDEPQRRSARLSAKPAPPKPEPKPKKAPAKK provided as SEQ ID NO: 24446) which is 31-amino acid fragment of HMGN2 protein that binds to nucleolin which is often found on the cell surface oof tumor and angiogenic endothelial cells.
  • the targeting peptide is CGKRK (provided as SEQ ID NO: 24447) which may target the angiogenic vessels of tumors.
  • the targeting peptide is a tumor-penetrating peptide with the CendR Motif such as, but not limited to, LyP-1 (CGNKRTRGC, with the CendR motif underlined and the sequence provided as SEQ ID NO: 24448), iRGD (CRGDKGPDC, with the CendR motif underlined and the sequence provided as SEQ ID NO: 24449), and iNGR (CRNGRGPDC, with the CendR motif underlined and the sequence provided as SEQ ID NO: 24450).
  • the CendR Motif such as, but not limited to, LyP-1 (CGNKRTRGC, with the CendR motif underlined and the sequence provided as SEQ ID NO: 24448), iRGD (CRGDKGPDC, with the CendR motif underlined and the sequence provided as SEQ ID NO: 2444
  • the targeting peptide is a T7 peptide (HAIYPRH, provided as SEQ ID NO: 24451), a MMP2-cleavable peptide (GPLGIAGQ, provided as SEQ ID NO: 24452), a CP15 peptide (VHLGYAT, provided as SEQ ID NO: 24453), a FSH peptide (YTRDLVYKDPARPKIQKTCTF, provided as SEQ ID NO: 24454), gastrin-releasing peptides (GRPs) (CGGNHWAVGHLM, provided as SEQ ID NO: 24455), and a RVG-brain delivery peptide (YTIWMPENPRPGTPCDIFTNSRGKRASNG, provided as SEQ ID NO: 24456).
  • HAIYPRH T7 peptide
  • GPLGIAGQ MMP2-cleavable peptide
  • VHLGYAT provided as SEQ ID NO: 24453
  • FSH peptide YTRDLVY
  • the ligand is a high molecular weight endogenous ligand such as transferrin (transferrin receptor is usually upregulated in many tumor cells), hyaluronic acid (HA) (negatively charged natural polymer which can bind to the surface CD44 receptors which are often overexpressed in primary and metastatic tumor cells), APOA1 (can bind with scavenger receptor class B-type 1 (SR-B1) which is found on the surface of hepatocytes).
  • the ligand is an aptamer, which is a small molecular weight (8-13 Kda) single-stranded RNA or DNA with low nanomolar binding affinities toward their targets.
  • the ligand is an antibody.
  • the ligand is a small molecule ligand such as, but not limited to, folate, anisamide, and galactose. (Leng et al. Journal of Drug Delivery. Vo. 17, Article ID 6971297; the contents of which is herein incorporated by reference in its entirety).
  • a hollow lipid core comprising a middle PLGA layer and an outer neutral lipid layer containing PEG may be used to delivery of the nucleic acid-based compositions of the present disclosure.
  • the lipid-polymer-lipid hybrid nanoparticles may comprise a core of the nucleic acid-based compositions disclosed herein and a polymer shell.
  • the polymer shell may be any of the polymers described herein and are known in the art.
  • the polymer shell may be used to protect the modified nucleic acids in the core.
  • Core–shell nanoparticles for use with the nucleic acid-based compositions of the present disclosure may be formed by the methods described in U.S. Pat. No. 8,313,777 herein incorporated by reference in its entirety.
  • the core-shell nanoparticles may comprise a core of the nucleic acid-based compositions disclosed herein and a polymer shell.
  • the polymer shell may be any of the polymers described herein and are known in the art.
  • the polymer shell may be used to protect the nucleic acid-based compositions in the core.
  • the core-shell nanoparticle may be used to treat an eye disease or disorder (See e.g.
  • the polymer used with the formulations described herein may be a modified polymer (such as, but not limited to, a modified polyacetal) as described in International Publication No. WO2011120053, herein incorporated by reference in its entirety.
  • Delivery [0285] The present disclosure encompasses the delivery of nucleic acid-based compositions including, for example, ASOs for any therapeutic, prophylactic, pharmaceutical, diagnostic or imaging use by any appropriate route taking into consideration likely advances in the sciences of drug delivery. Delivery may be naked or formulated.
  • the nucleic acid-based compositions of the present disclosure may be delivered to a cell naked.
  • naked refers to delivering nucleic acid-based compositions free from agents which promote transfection.
  • the nucleic acid-based compositions delivered to the cell may contain no modifications.
  • the naked nucleic acid-based compositions may be delivered to the cell using routes of administration known in the art and described herein.
  • the nucleic acid-based compositions of the present disclosure may be formulated, using the methods described herein.
  • the formulations may contain nucleic acid-based compositions which may be modified and/or unmodified.
  • the formulations may further include, but are not limited to, cell penetration agents, a pharmaceutically acceptable carrier, a delivery agent, a bioerodible or biocompatible polymer, a solvent, and a sustained-release delivery depot.
  • the formulated nucleic acid-based compositions may be delivered to the cell using routes of administration known in the art and described herein.
  • the compositions may also be formulated for direct delivery to an organ or tissue in any of several ways in the art including, but not limited to, direct soaking or bathing, via a catheter, by gels, powder, ointments, creams, gels, lotions, and/or drops, by using substrates such as fabric or biodegradable materials coated or impregnated with the compositions, and the like.
  • the nucleic acid-based compositions of the present disclosure may also be cloned into a retroviral replicating vector (RRV) and transduced to cells.
  • RRV retroviral replicating vector
  • the nucleic acid-based compositions of the present disclosure may be administered by any route which results in a therapeutically effective outcome. These include, but are not limited to enteral, gastroenteral, epidural, oral, transdermal, epidural (peridural), intracerebral (into the cerebrum), intracerebroventricular (into the cerebral ventricles), epicutaneous (application onto the skin), intradermal, (into the skin itself), subcutaneous (under the skin), nasal administration (through the nose), intravenous (into a vein), intraarterial (into an artery), intramuscular (into a muscle), intracardiac (into the heart), intraosseous infusion (into the bone marrow), intrathecal (into the spinal canal or into the subarachnoid space to reach the CSF), intraperitoneal, (infusion or injection into the peritoneum), intravesical infusion (e.g., into the bladder using a catheter), intravitreal, (through the eye), intracaver
  • compositions may be administered in a way which allows them to cross the blood-brain barrier, vascular barrier, or other epithelial barrier.
  • Routes of administration disclosed in International Publication WO 2013/090648 filed December 14, 2012, the contents of which are incorporated herein by reference in their entirety, may be used to administer the nucleic acid-based compositions of the present disclosure.
  • Delivery of modified therapeutic compounds described herein to a subject over prolonged periods of time, for example, for periods of one week to one year, may be accomplished by a single administration of a controlled release system containing sufficient active ingredient for the desired release period.
  • controlled release systems such as monolithic or reservoir-type microcapsules, depot implants, polymeric hydrogels, osmotic pumps, vesicles, micelles, liposomes, transdermal patches, iontophoretic devices and alternative injectable dosage forms may be utilized for this purpose. Localization at the site to which delivery of the active ingredient is desired is an additional feature of some controlled release devices, which may prove beneficial in the treatment of certain disorders.
  • delivery across the barrier of the skin would be enhanced using electrodes (e.g. iontophoresis), electroporation, or the application of short, high-voltage electrical pulses to the skin, radiofrequencies, ultrasound (e.g.
  • the drug can be included in single-layer drug-in-adhesive, multi-layer drug-in-adhesive, reservoir, matrix, or vapor style patches, or could utilize patchless technology. Delivery across the barrier of the skin could also be enhanced using encapsulation, a skin lipid fluidizer, or a hollow or solid microstructured transdermal system (MTS, such as that manufactured by 3M), jet injectors.
  • MTS microstructured transdermal system
  • Additives to the formulation to aid in the passage of therapeutic compounds through the skin include prodrugs, chemicals, surfactants, cell penetrating peptides, permeation enhancers, encapsulation technologies, enzymes, enzyme inhibitors, gels, nanoparticles and peptide or protein chaperones.
  • One form of controlled-release formulation contains the therapeutic compound or its salt dispersed or encapsulated in a slowly degrading, non-toxic, non-antigenic polymer such as copoly(lactic/glycolic) acid, as described in the pioneering work of Kent et al., US Patent No. 4,675,189, incorporated by reference herein.
  • the compounds, or their salts may also be formulated in cholesterol or other lipid matrix pellets, or silastomer matrix implants. Additional slow release, depot implant or injectable formulations will be apparent to the skilled artisan. See, for example, Sustained and Controlled Release Drug Delivery Systems, JR Robinson ed., Marcel Dekker Inc., New York, 1978; and Controlled Release of Biologically Active Agents, RW Baker, John Wiley & Sons, New York, 1987. The foregoing are incorporated by reference in their entirety.
  • An additional form of controlled-release formulation comprises a solution of biodegradable polymer, such as copoly(lactic/glycolic acid) or block copolymers of lactic acid and PEG, is a bioacceptable solvent, which is injected subcutaneously or intramuscularly to achieve a depot formulation. Mixing of the therapeutic compounds described herein with such a polymeric formulation is suitable to achieve very long duration of action formulations.
  • biodegradable polymer such as copoly(lactic/glycolic acid) or block copolymers of lactic acid and PEG
  • the absorption across the nasal mucous membrane may be further enhanced by surfactants, such as, for example, glycocholic acid, cholic acid, taurocholic acid, ethocholic acid, deoxycholic acid, chenodeoxycholic acid, dehdryocholic acid, glycodeoxycholic acid, cycledextrins and the like in an amount in the range of between about 0.1 and 15 weight percent, between about 0.5 and 4 weight percent, or about 2 weight percent.
  • surfactants such as, for example, glycocholic acid, cholic acid, taurocholic acid, ethocholic acid, deoxycholic acid, chenodeoxycholic acid, dehdryocholic acid, glycodeoxycholic acid, cycledextrins and the like in an amount in the range of between about 0.1 and 15 weight percent, between about 0.5 and 4 weight percent, or about 2 weight percent.
  • compositions may be in the form of a sterile injectable preparation, for example, as a sterile injectable aqueous or oleaginous suspension.
  • This suspension may be formulated according to techniques known in the art using suitable dispersing or wetting agents (such as, for example, Tween 80) and suspending agents.
  • the sterile injectable preparation may also be a sterile injectable solution or suspension in a non- toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1,3-butanediol.
  • a non- toxic parenterally-acceptable diluent or solvent for example, as a solution in 1,3-butanediol.
  • acceptable vehicles and solvents that may be employed are mannitol, water, Ringer's solution and isotonic sodium chloride solution.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • any bland fixed oil may be employed including synthetic mono- or diglycerides.
  • Fatty acids such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions. These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant such as Ph. Helv or a similar alcohol.
  • the pharmaceutical compositions of the present disclosure may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, and aqueous suspensions and solutions. In the case of tablets for oral use, carriers that are commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added.
  • compositions of present disclosure may also be administered in the form of suppositories for rectal administration.
  • a suitable non-irritating excipient that is solid at room temperature but liquid at the rectal temperature and therefore will melt in the rectum to release the active components.
  • suitable non-irritating excipient include, but are not limited to, cocoa butter, beeswax and polyethylene glycols.
  • Topical administration of the pharmaceutical compositions of the present disclosure is especially useful when the desired treatment involves areas or organs readily accessible by topical application.
  • the pharmaceutical composition should be formulated with a suitable ointment containing the active components suspended or dissolved in a carrier.
  • Carriers for topical administration of the compounds of the present disclosure include, but are not limited to, mineral oil, liquid petroleum, white petroleum, propylene glycol, polyoxyethylene polyoxypropylene compound, emulsifying wax and water.
  • the pharmaceutical composition can be formulated with a suitable lotion or cream containing the active compound suspended or dissolved in a carrier.
  • Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.
  • the pharmaceutical compositions of the present disclosure may also be topically applied to the lower intestinal tract by rectal suppository formulation or in a suitable enema formulation. Topical transdermal patches are also included in the present disclosure.
  • the pharmaceutical compositions of the present disclosure may be administered by nasal aerosol or inhalation.
  • compositions are prepared according to techniques well- known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other solubilizing or dispersing agents known in the art.
  • a number of formulations offer advantages. Adsorption of the therapeutic compound to readily dispersed solids such as diketopiperazines (for example, Technosphere particles (Pfutzner, A and Forst, T, 2005, Expert Opin Drug Deliv 2:1097-1106) or similar structures gives a formulation that results in rapid initial uptake of the therapeutic compound.
  • Lyophilized powders, especially glassy particles, containing the therapeutic compound and an excipient are useful for delivery to the lung with good bioavailability, for example, see Exubera ® (inhaled insulin, Pfizer, Inc. and Aventis Pharmaceuticals Inc.) and Afrezza ® (inhaled insulin, Mannkind, Corp.).
  • the pharmaceutical compositions of the present disclosure may be administered by local delivery to the bladder such as, but not limited to, intravesical therapy.
  • Such compositions are prepared according to techniques well-known in the art of pharmaceutical formulation and may be prepared as solutions that can be delivered using, for example, a catheter that is put into the bladder through the urethra.
  • compositions of the present disclosure may be formulated to be administered to the CNS by routes known in the art such as, but not limited to, direct intraparenchymal administration, intrathecal delivery and intracerebroventricular infusion.
  • the pharmaceutical compositions are formulated to have the biodistribution of the pharmaceutical composition located in the tumor cells.
  • the pharmaceutical compositions of the present disclosure may be formulated to improve delivery to tumors.
  • a pharmaceutical composition described herein can be formulated into a dosage form described herein, such as a topical, intranasal, intratracheal, or injectable (e.g., intravenous, intraocular, intravitreal, intramuscular, intracardiac, intraperitoneal, subcutaneous).
  • injectable e.g., intravenous, intraocular, intravitreal, intramuscular, intracardiac, intraperitoneal, subcutaneous.
  • Liquid dosage forms, injectable preparations, pulmonary forms, and solid dosage forms described in International Publication WO 2013/090648 filed December 14, 2012, the contents of which are incorporated herein by reference in their entirety may be used as dosage forms for the nucleic acid-based compositions of the present disclosure.
  • the present disclosure contemplates dosage levels of between about 0.001 and about 100 mg GABPB1-hybridizing oligonucleotide (e.g., GABPB1-targeting ASO)/kg body weight per day, preferably between about 0.005 and about 50 mg/kg, 0.01 and about 10 mg/kg, 0.05 and about 5 mg/kg, 0.1 and about 1 mg/kg body weight.
  • Other embodiments contemplate a dosage of between about 0.001-0.010, 0.010-0.050, 0.050-0.100, 0.1-0.5, 0.5- 1.0, 1.0-5.0, 5.0-10, or 10-50 mg/kg body weight.
  • the dosages may be administered about hourly, multiple times per day, daily, every other day, weekly, every other week, monthly, or on an as-needed basis.
  • Such administration can be used as a chronic or acute therapy.
  • the amount of drug that may be combined with the carrier to produce a single dosage form will vary depending upon the host treated and the particular mode of administration.
  • a typical preparation will contain from about 5% to about 95% active compound (w/w).
  • such preparations contain from about 20% to about 80%, 30% to about 70%, 40% to about 60%, or about 50% active compound.
  • the preparations used in the present disclosure will be about 5-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90- 99%, or greater than 99% of the active ingredient.
  • a maintenance dose of a compound, composition or combination of the present disclosure may be administered, if necessary.
  • the dosage or frequency of administration, or both may be reduced, as a function of the symptoms, to a level at which the improved condition is retained when the symptoms have been alleviated to the desired level, treatment should cease.
  • Patients may, however, require intermittent treatment on a long-term basis upon any recurrence of disease symptoms.
  • nucleic acid-based compositions of the present disclosure provides methods of using nucleic acid-based compositions of the present disclosure and pharmaceutical compositions comprising the nucleic acid-based compositions and at least one pharmaceutically acceptable carrier.
  • the nucleic acid-based compositions of the present disclosure modulate the expression of the target gene, i.e., GABPB1.
  • a method of regulating the expression of a target gene in vitro and/or in vivo comprising administering the nucleic acid- based compositions of the present disclosure.
  • the expression of the target gene is decreased by at least 5%, 10%, 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the nucleic acid-based compositions of the present disclosure compared to the expression of the target gene in the absence of the nucleic acid-based compositions of the present disclosure.
  • the expression of the target gene is decreased by a factor of at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or by a factor of at least 15, 20, 25, 30, 35, 40, 45, 50, or by a factor of at least 60, 70, 80, 90, 100, in the presence of the nucleic acid-based compositions of the present disclosure compared to the expression of the target gene in the absence of the nucleic acid-based compositions of the present disclosure.
  • One aspect of the present application provides a method of modulating the expression of GABPB1 gene comprising administering GABPB1-hybridizing oligonucleotides of the present disclosure.
  • the expression of GABPB1 gene is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure compared to the expression of GABPB1 gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure.
  • the expression of GABPB1 gene is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure which specifically targets the GABPB1L isoform as compared to the expression of GABPB1 gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure specifically targeting GABPB1L.
  • the expression of GABPB1 gene is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1- hybridizing oligonucleotides of the present disclosure which specifically targets the GABPB1S isoform as compared to the expression of GABPB1 gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure specifically targeting GABPB1S.
  • the expression of GABPB1 gene is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure which specifically targets both the GABPB1L and GABPB1S isoforms (total GABPB1) as compared to the expression of GABPB1 gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure specifically targeting total GABPB1.
  • the expression of GABPB1 gene is decreased by a factor of at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or by a factor of at least 15, 20, 25, 30, 35, 40, 45, 50, or by a factor of at least 60, 70, 80, 90, 100, in the presence of the GABPB1-hybridizing oligonucleotides (e.g., GABPB1-hybridizing oligonucleotides targeting the GABPB1L isoform, the GABPB1S isoform or total GABPB1) of the present disclosure compared to the expression of GABPB1 gene in the absence of the GABP-hybridizing oligonucleotides of the present disclosure.
  • the GABPB1-hybridizing oligonucleotides e.g., GABPB1-hybridizing oligonucleotides targeting the GABPB1L isoform, the GABPB1S isoform or total GABPB1
  • the expression of TERT is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure compared to the expression of TERT in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure.
  • the expression of TERT gene is decreased by a factor of at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or by a factor of at least 15, 20, 25, 30, 35, 40, 45, 50, or by a factor of at least 60, 70, 80, 90, 100, in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure compared to the expression of TERT gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure.
  • the modulation of the expression of GABPB1 gene may be reflected in or determined by a change in TERT protein levels.
  • Some embodiments provide methods of use of the nucleic acid-based compositions described herein to prevent or treat diseases or disorders associated with telomerase expression such as, but not limited to cancer.
  • the methods provided herein include administering GABPB1-hybridizing oligonucleotides described herein to subjects having a cancer.
  • the methods of administering the GABPB1-hybridizing oligonucleotides decrease expression of the GABPB1 gene, and, in some cases, decrease the level of TERT mRNA expression in cancer cells harboring TERT promoter mutations which may ultimately reduce the expression of telomerase in the subject. Accordingly, the present disclosure provides methods for treating individual subjects suffering from cancer.
  • the methods of use can be assessed using any endpoint indicating a benefit to the subject, including, without limitation, (1) inhibition, to some extent, of disease progression, including stabilization, slowing down and complete arrest; (2) reduction in the number of disease episodes and/or symptoms; (3) inhibition (i.e., reduction, slowing down or complete stopping) of a disease cell infiltration into adjacent peripheral organs and/or tissues; (4) inhibition (i.e.
  • cancer refers to any of various malignant neoplasms characterized by the proliferation of anaplastic cells that tend to invade surrounding tissue and metastasize to new body sites and also refers to the pathological condition characterized by such malignant neoplastic growths.
  • Cancers may be tumors or hematological malignancies, and include but are not limited to, all types of lymphomas/leukemias, carcinomas and sarcomas, such as those cancers or tumors found in the anus, bladder, bile duct, bone, brain, breast, cervix, colon/rectum, endometrium, esophagus, eye, gallbladder, head and neck, liver, kidney, larynx, lung, mediastinum (chest), mouth, ovaries, pancreas, penis, prostate, skin, small intestine, stomach, spinal marrow, tailbone, testicles, thyroid and uterus.
  • lymphomas/leukemias such as those cancers or tumors found in the anus, bladder, bile duct, bone, brain, breast, cervix, colon/rectum, endometrium, esophagus, eye, gallbladder, head and neck, liver, kidney, larynx, lung, mediastinum (ches
  • Types of carcinomas which may be treated with the compositions of the present disclosure include, but are not limited to, papilloma/carcinoma, choriocarcinoma, endodermal sinus tumor, teratoma, adenoma/adenocarcinoma, melanoma, fibroma, lipoma, leiomyoma, rhabdomyoma, mesothelioma, angioma, osteoma, chondroma, glioma, lymphoma/leukemia, squamous cell carcinoma, small cell carcinoma, large cell undifferentiated carcinomas, basal cell carcinoma and sinonasal undifferentiated carcinoma.
  • Types of carcinomas which may be treated with the compositions of the present disclosure include, but are not limited to, soft tissue sarcoma such as alveolar soft part sarcoma, angiosarcoma, dermatofibrosarcoma, desmoid tumor, desmoplastic small round cell tumor, extraskeletal chondrosarcoma, extraskeletal osteosarcoma, fibrosarcoma, hemangiopericytoma, hemangiosarcoma, Kaposi's sarcoma, leiomyosarcoma, liposarcoma, lymphangiosarcoma, lymphosarcoma, malignant fibrous histiocytoma, neurofibrosarcoma, rhabdomyosarcoma, synovial sarcoma, and Askin's tumor, Ewing's sarcoma (primitive neuroectodermal tumor), malignant hemangioendothelioma, malignant schwannoma, osteosar
  • the carcinoma which may be treated may be Acute granulocytic leukemia, Acute lymphocytic leukemia, Acute myelogenous leukemia, Adenocarcinoma, Adenosarcoma, Adrenal cancer, Adrenocortical carcinoma, Anal cancer, Anaplastic astrocytoma, Angiosarcoma, Appendix cancer, Astrocytoma, Basal cell carcinoma, B-Cell lymphoma ), Bile duct cancer, Bladder cancer, Bone cancer, Bowel cancer, Brain cancer, Brain stem glioma, Brain tumor, Breast cancer, Carcinoid tumors, Cervical cancer, Cholangiocarcinoma, Chondrosarcoma, Chronic lymphocytic leukemia, Chronic myelogenous leukemia, Colon cancer, Colorectal cancer, Craniopharyngioma, Cutaneous lymphoma, Cutaneous melanoma, Diffuse astro
  • kits and Devices Kits The disclosure provides a variety of kits for conveniently and/or effectively carrying out methods of the present disclosure. Typically, kits will comprise sufficient amounts and/or numbers of components to allow a user to perform multiple treatments of a subject(s) and/or to perform multiple experiments. [0319] In some embodiments, the present disclosure provides kits for modulating the expression of genes in vitro or in vivo, comprising nucleic acid-based compositions of the present disclosure or a combination of nucleic acid-based compositions of the present disclosure, nucleic acid-based compositions modulating other genes, siRNAs, miRNAs or other oligonucleotide molecules.
  • the kit may further comprise packaging and instructions and/or a delivery agent to form a formulation, e.g., for administration to a subject in need of treatment using the nucleic acid-based compositions described herein.
  • the delivery agent may comprise a saline, a buffered solution, a lipidoid, a dendrimer or any suitable delivery agent.
  • the buffer solution may include sodium chloride, calcium chloride, phosphate and/or EDTA.
  • the buffer solution may include, but is not limited to, saline, saline with 2mM calcium, 5% sucrose, 5% sucrose with 2mM calcium, 5% Mannitol, 5% Mannitol with 2mM calcium, Ringer’s lactate, sodium chloride, sodium chloride with 2mM calcium and mannose (See U.S. Pub. No. 20120258046; herein incorporated by reference in its entirety).
  • the buffer solutions may be precipitated or it may be lyophilized. The amount of each component may be varied to enable consistent, reproducible higher concentration saline or simple buffer formulations.
  • the components may also be varied in order to increase the stability of nucleic acid-based compositions in the buffer solution over a period of time and/or under a variety of conditions.
  • Devices [0322] The present disclosure provides for devices which may incorporate nucleic acid- based compositions of the present disclosure. These devices can contain a stable formulation available to be immediately delivered to a subject in need thereof, such as a human patient.
  • Non-limiting examples of the devices include a pump, a catheter, a needle, a transdermal patch, a pressurized olfactory delivery device, electroporation devices, iontophoresis devices, multi-layered microfluidic devices.
  • the devices may be employed to deliver nucleic acid-based compositions of the present disclosure according to single, multi- or split-dosing regiments.
  • the devices may be employed to deliver nucleic acid-based compositions of the present disclosure across biological tissue, intradermal, subcutaneously, or intramuscularly. More examples of devices suitable for delivering oligonucleotides are disclosed in International Publication WO 2013/090648, the contents of which are incorporated herein by reference in their entirety. V. Definitions [0324] For convenience, the meaning of certain terms and phrases used in the specification, examples, and appended claims, are provided below. If there is an apparent discrepancy between the usage of a term in other parts of this specification and its definition provided in this section, the definition in this section shall prevail.
  • Administered in combination means that two or more agents are administered to a subject at the same time or within an interval such that there may be an overlap of an effect of each agent on the patient. In some embodiments, they are administered within about 60, 30, 15, 10, 5, or 1 minute of one another. In some embodiments, the administrations of the agents are spaced sufficiently close together such that a combinatorial (e.g., a synergistic) effect is achieved.
  • Agonist refers to a substance that binds to a receptor and activates the receptor to produce a biological response. They can be in the form of antibodies, antigen-binding fragments, proteins, peptides, glycoproteins, glycopeptides, glycolipids, polysaccharides, oligosaccharides, nucleic acids, bioorganic molecules, peptidomimetics, chemicals, pharmacological agents and their metabolites, and the like.
  • an "antagonist” refers to a molecule capable of neutralizing, blocking, inhibiting, abrogating, reducing or interfering with the activities of a particular or specified protein, including its binding to one or more receptors in the case of a ligand, or binding to one or more ligands in case of a receptor.
  • Amino acid As used herein, the terms “amino acid” and “amino acids” refer to all naturally occurring L-alpha-amino acids.
  • amino acids are identified by either the one- letter or three-letter designations as follows: aspartic acid (Asp:D), isoleucine (Ile:I), threonine (Thr:T), leucine (Leu:L), serine (Ser:S), tyrosine (Tyr:Y), glutamic acid (Glu:E), phenylalanine (Phe:F), proline (Pro:P), histidine (His:H), glycine (Gly:G), lysine (Lys:K), alanine (Ala:A), arginine (Arg:R), cysteine (Cys:C), tryptophan (Trp:W), valine (Val:V), glutamine (Gln:Q) methionine (Met:M), asparagines (Asn:N), where the amino acid is listed first followed parenthetically by the three and one letter codes, respectively.
  • animal refers to any member of the animal kingdom. In some embodiments, “animal” refers to humans at any stage of development. In some embodiments, “animal” refers to non-human animals at any stage of development. In certain embodiments, the non-human animal is a mammal (e.g., a rodent, a mouse, a rat, a rabbit, a monkey, a dog, a cat, a sheep, cattle, a primate, or a pig). In some embodiments, animals include, but are not limited to, mammals, birds, reptiles, amphibians, fish, and worms.
  • mammals include, but are not limited to, mammals, birds, reptiles, amphibians, fish, and worms.
  • the animal is a transgenic animal, genetically-engineered animal, or a clone.
  • Antisense oligonucleotide (ASO) “Antisense oligonucleotides” or “ASOs” are typically short (e.g., around 8 – 50 nucleotides), single-stranded oligonucleotide molecules designed to hybridize with an RNA sequence, e.g., mRNA, rRNA. Such targeting may result in either an upregulation or downregulation of the expression of the gene depending on the design of the ASO and the site of action, or localization of the ASO.
  • ASOs include oligodeoxyribonucleotide or oligoribonucleotide molecules that downregulate or have a negative effect on the expression of a specific gene.
  • the ASOs can be between 8 to 20 nucleotides in length.
  • the ASOs are designed to complementarily bind to a region of a target gene RNA, including pre-mRNA or mRNA, the target gene being GABPB1.
  • GABPB1-ASO an ASO that downregulates the expression of the GABPB1 gene.
  • ASOs as described herein include ASOs having modified nucleotides, including fully modified and partially modified ASOs.
  • Gapmer ASOs are short single- stranded ASOs containing a central DNA sequence commonly flanked by a locked nucleic acid (LNA) sequence that interrupts mRNA expression by induction of RNase H activation. They can exhibit cellular entry without the necessity of a transfection agent by a process termed gymnosis. [0331] Approximately: As used herein, the term “approximately” or “about,” as applied to one or more values of interest, refers to a value that is similar to a stated reference value.
  • LNA locked nucleic acid
  • the term “approximately” or “about” refers to a range of values that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value).
  • association means that the moieties are physically associated or connected with one another, either directly or via one or more additional moieties that serves as a linking agent, to form a structure that is sufficiently stable so that the moieties remain physically associated under the conditions in which the structure is used, e.g., physiological conditions.
  • An “association” need not be strictly through direct covalent chemical bonding. It may also suggest ionic or hydrogen bonding or a hybridization based connectivity sufficiently stable such that the “associated” entities remain physically associated.
  • Bifunction or Bifunctional refers to any substance, molecule or moiety which is capable of or maintains at least two functions. The functions may affect the same outcome or a different outcome. The structure that produces the function may be the same or different.
  • bifunctional nucleic acid-based compositions of the present disclosure may comprise a cytotoxic peptide (a first function) while those nucleosides which comprise the nucleic acid-based compositions are, in and of themselves, cytotoxic (second function).
  • Biocompatible As used herein, the term “biocompatible” means compatible with living cells, tissues, organs or systems posing little to no risk of injury, toxicity or rejection by the immune system.
  • Biodegradable As used herein, the term “biodegradable” means capable of being broken down into innocuous products by the action of living things.
  • Biologically active As used herein, the phrase “biologically active” refers to a characteristic of any substance that has activity in a biological system and/or organism. For instance, a substance that, when administered to an organism, has a biological effect on that organism, is considered to be biologically active.
  • the nucleic acid- based compositions of the present disclosure may be considered biologically active if even a portion of the nucleic acid-based compositions is biologically active or mimics an activity considered biologically relevant.
  • cancer in an individual refers to the presence of cells possessing characteristics typical of cancer-causing cells, such as uncontrolled proliferation, immortality, metastatic potential, rapid growth and proliferation rate, and certain characteristic morphological features. Often, cancer cells will be in the form of a tumor, but such cells may exist alone within an individual, or may circulate in the blood stream as independent cells, such as leukemic cells.
  • Cell growth As used herein, the term “cell growth” is principally associated with growth in cell numbers, which occurs by means of cell reproduction (i.e. proliferation) when the rate of the latter is greater than the rate of cell death (e.g.
  • Cell type refers to a cell from a given source (e.g., a tissue, organ) or a cell in a given state of differentiation, or a cell associated with a given pathology or genetic makeup.
  • Chromosome As used herein, the term “chromosome” refers to an organized structure of DNA and protein found in cells.
  • Complementary As used herein, the term “complementary” as it relates to nucleic acids refers to hybridization or base pairing between nucleotides or nucleic acids, such as, for example, between the two strands of a double-stranded DNA molecule or between an oligonucleotide probe and a target.
  • Condition As used herein, the term “condition” refers to the status of any cell, organ, organ system or organism. Conditions may reflect a disease state or simply the physiologic presentation or situation of an entity.
  • Controlled Release refers to a pharmaceutical composition or compound release profile that conforms to a particular pattern of release to effect a therapeutic outcome.
  • Cytotoxic refers to killing or causing injurious, toxic, or deadly effect on a cell (e.g., a mammalian cell (e.g., a human cell)), bacterium, virus, fungus, protozoan, parasite, prion, or a combination thereof.
  • Delivery refers to the act or manner of delivering a compound, substance, entity, moiety, cargo or payload.
  • Delivery Agent refers to any substance which facilitates, at least in part, the in vivo delivery of nucleic acid-based compositions of the present disclosure to targeted cells.
  • Destabilized As used herein, the term “destable,” “destabilize,” or “destabilizing region” means a region or molecule that is less stable than a starting, wild-type or native form of the same region or molecule.
  • Detectable label refers to one or more markers, signals, or moieties which are attached, incorporated or associated with another entity that is readily detected by methods known in the art including radiography, fluorescence, chemiluminescence, enzymatic activity, absorbance and the like. Detectable labels include radioisotopes, fluorophores, chromophores, enzymes, dyes, metal ions, ligands such as biotin, avidin, streptavidin and haptens, quantum dots, and the like. Detectable labels may be located at any position in the oligonucleotides disclosed herein. They may be within the nucleotides or located at the 5’ or 3’ terminus.
  • Downregulation refers to a decrease in the level of expression of the gene, a decrease in the level of the polypeptide(s) encoded by the gene, the level bioactive gene product, or level of the RNA transcript(s) transcribed from the template strand of the gene as compared to the levels observed in the absence of the nucleic acid-based composition(s) described herein.
  • the nucleic acid-based compositions of the present disclosure may have a direct downregulating effect on the expression of the target gene.
  • the term “downregulation,” depending on the context, can also refer to the effect of a trans-acting activator of expression.
  • Downregulatory sequence element The terms “DSE” or “downregulatory sequence element” as used in the context of expression control elements in a gene or a transcript of a gene, including a target transcript, refer to those cis-acting sequence-based or secondary structure-based features of a sequence which, under normal physiological conditions, tend to decrease or inhibit translation of the transcript and/or expression of the gene.
  • the term “downregulation,” depending on the context, can also refer to the effect of a trans-acting inhibitor of expression.
  • Effective Amount As used herein, an "effective amount” refers to an amount of therapeutic compound that is effective, at dosages and for periods of time necessary, to achieve the desired therapeutic or prophylactic result.
  • Element refers to a particular feature of the sequence, such as a known binding site for a protein or other factor or a region that is involved in the formation of secondary structure of the nucleotide sequence. Elements include, for example, upstream open reading frames (uORFs), internal ribosome entry sites (IRES), upstream initiation codons, upstream termination codons, and binding sites for proteins and other factors that operate in trans with respect to the nucleotide sequence.
  • uORFs upstream open reading frames
  • IRS internal ribosome entry sites
  • Encapsulate means to enclose, surround or encase.
  • Engineered As used herein, embodiments of the disclosure are “engineered” when they are designed to have a feature or property, whether structural or chemical, that varies from a starting point, wild type or native molecule.
  • Equivalent subject As used herein, “equivalent subject” may be e.g. a subject of similar age, sex and health such as liver health or cancer stage, or the same subject prior to treatment according to the disclosure. The equivalent subject is "untreated” in that he does not receive treatment with nucleic acid-based compositions according to the disclosure. However, he may receive a conventional anti-cancer treatment, provided that the subject who is treated with the nucleic acid-based compositions of the disclosure receives the same or equivalent conventional anti-cancer treatment.
  • Exosome As used herein, “exosome” is a vesicle secreted by mammalian cells.
  • Expression As used herein, “expression” of a nucleic acid sequence refers to one or more of the following events: (1) production of an RNA template from a DNA sequence (e.g., by transcription); (2) processing of an RNA transcript (e.g., by splicing, editing, 5’ cap formation, and/or 3’ end processing); (3) translation of an RNA into a polypeptide or protein; and (4) post-translational modification of a polypeptide or protein.
  • Feature As used herein, a “feature” refers to a characteristic, a property, or a distinctive element.
  • a “formulation” includes at least one nucleic acid- based composition of the present disclosure and a delivery agent.
  • Fragment A “fragment,” as used herein, refers to a portion.
  • fragments of proteins may comprise polypeptides obtained by digesting full-length protein isolated from cultured cells.
  • Fragments of oligonucleotides may comprise nucleotides, or regions of nucleotides.
  • Functional As used herein, a “functional” biological molecule is a biological molecule in a form in which it exhibits a property and/or activity by which it is characterized.
  • Gene refers to a nucleic acid sequence that comprises control and most often coding sequences necessary for producing a polypeptide or precursor. Genes, however, may not be translated and instead code for regulatory or structural RNA molecules.
  • a gene may be derived in whole or in part from any source known to the art, including a plant, a fungus, an animal, a bacterial genome or episome, eukaryotic, nuclear or plasmid DNA, cDNA, viral DNA, or chemically synthesized DNA.
  • a gene may contain one or more modifications in either the coding or the untranslated regions that could affect the biological activity or the chemical structure of the expression product, the rate of expression, or the manner of expression control.
  • Gene expression refers to the process by which a nucleic acid sequence undergoes successful transcription and in most instances translation to produce a protein or peptide.
  • RNA or mRNA the nucleic acid product of transcription
  • amino acid product of translation e.g., polypeptides or peptides.
  • Genome The term “genome” is intended to include the entire DNA complement of an organism, including the nuclear DNA component, chromosomal or extrachromosomal DNA, as well as the cytoplasmic domain (e.g., mitochondrial DNA).
  • homolog As used herein, the term “homologs” are bioactive molecules that are similar to a reference molecule at the nucleotide sequence, peptide sequence, functional, or structural level. Homologs may include sequence derivatives that share a certain percent identity with the reference sequence.
  • Homologous or derivative nucleic acid sequences may also be defined by their ability to remain bound to a reference nucleic acid sequence under high stringency hybridization conditions. Homologs having a structural or functional similarity to a reference molecule may be chemical derivatives of the reference molecule. [0367] Homology: As used herein, the term “homology” refers to the overall relatedness between polymeric molecules, e.g. between nucleic acid molecules (e.g. DNA molecules and/or RNA molecules) and/or between polypeptide molecules.
  • polymeric molecules are considered to be “homologous” to one another if their sequences are at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical or similar.
  • the term “homologous” necessarily refers to a comparison between at least two sequences (polynucleotide or polypeptide sequences).
  • two polynucleotide sequences are considered to be homologous if the polypeptides they encode are at least about 50%, 60%, 70%, 80%, 90%, 95%, or even 99% for at least one stretch of at least about 20 amino acids.
  • homologous polynucleotide sequences are characterized by the ability to encode a stretch of at least 4–5 uniquely specified amino acids. For polynucleotide sequences less than 60 nucleotides in length, homology is determined by the ability to encode a stretch of at least 4–5 uniquely specified amino acids. In accordance with the disclosure, two protein sequences are considered to be homologous if the proteins are at least about 50%, 60%, 70%, 80%, or 90% identical for at least one stretch of at least about 20 amino acids. [0368] Identity: As used herein, the term “identity” refers to the overall relatedness between polymeric molecules, e.g., between oligonucleotide molecules (e.g.
  • DNA molecules and/or RNA molecules DNA molecules and/or RNA molecules) and/or between polypeptide molecules.
  • Calculation of the percent identity of two polynucleotide sequences can be performed by aligning the two sequences for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second nucleic acid sequences for optimal alignment and non-identical sequences can be disregarded for comparison purposes).
  • the length of a sequence aligned for comparison purposes is at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or 100% of the length of the reference sequence.
  • the nucleotides at corresponding nucleotide positions are then compared.
  • the percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which needs to be introduced for optimal alignment of the two sequences.
  • the comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm.
  • the percent identity between two nucleotide sequences can be determined using methods such as those described in Computational Molecular Biology, Lesk, A. M., ed., Oxford University Press, New York, 1988; Biocomputing: Informatics and Genome Projects, Smith, D.
  • the percent identity between two nucleotide sequences can be determined using the algorithm of Meyers and Miller (CABIOS, 1989, 4:11-17), which has been incorporated into the ALIGN program (version 2.0) using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4.
  • the percent identity between two nucleotide sequences can, alternatively, be determined using the GAP program in the GCG software package using an NWSgapdna.CMP matrix. Methods commonly employed to determine percent identity between sequences include, but are not limited to those disclosed in Carillo, H., and Lipman, D., SIAM J Applied Math., 48:1073 (1988); incorporated herein by reference.
  • Inhibit expression of a gene means to cause a reduction in the amount of an expression product of the gene.
  • the expression product can be an RNA transcribed from the gene (e.g., an mRNA) or a polypeptide translated from an mRNA transcribed from the gene. Typically, a reduction in the level of an mRNA results in a reduction in the level of a polypeptide translated therefrom.
  • the level of expression may be determined using standard techniques for measuring mRNA or protein.
  • In vitro refers to events that occur in an artificial environment, e.g., in a test tube or reaction vessel, in cell culture, in a Petri dish, etc., rather than within an organism (e.g., animal, plant, or microbe).
  • Isolated refers to a substance or entity that has been separated from at least some of the components with which it was associated (whether in nature or in an experimental setting). Isolated substances may have varying levels of purity in reference to the substances from which they have been associated. Isolated substances and/or entities may be separated from at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or more of the other components with which they were initially associated.
  • isolated agents are more than about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% pure.
  • a substance is “pure” if it is substantially free of other components.
  • substantially isolated By “substantially isolated” is meant that the compound is substantially separated from the environment in which it was formed or detected. Partial separation can include, for example, a composition enriched in the compound of the present disclosure.
  • Substantial separation can include compositions containing at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% by weight of the compound of the present disclosure, or salt thereof. Methods for isolating compounds and their salts are routine in the art.
  • Label refers to a substance or a compound which is incorporated into an object so that the substance, compound or object may be detectable.
  • Linker refers to a group of atoms, e.g., 10-1,000 atoms, and can be comprised of the atoms or groups such as, but not limited to, carbon, amino, alkylamino, oxygen, sulfur, sulfoxide, sulfonyl, carbonyl, and imine.
  • the linker can be attached to a modified nucleoside or nucleotide on the nucleobase or sugar moiety at a first end, and to a payload, e.g., a detectable or therapeutic agent, at a second end.
  • the linker may be of sufficient length as to not interfere with incorporation into a nucleic acid sequence.
  • the linker can be used for any useful purpose, such as to form conjugates, as well as to administer a payload, as described herein.
  • Examples of chemical groups that can be incorporated into the linker include, but are not limited to, alkyl, alkenyl, alkynyl, amido, amino, ether, thioether, ester, alkylene, heteroalkylene, aryl, or heterocyclyl, each of which can be optionally substituted, as described herein.
  • linkers include, but are not limited to, unsaturated alkanes, polyethylene glycols (e.g., ethylene or propylene glycol monomeric units, e.g., diethylene glycol, dipropylene glycol, triethylene glycol, tripropylene glycol, tetraethylene glycol, or tetraethylene glycol), and dextran polymers and derivatives thereof.
  • Non-limiting examples of a selectively cleavable bond include an amido bond can be cleaved for example by the use of tris(2-carboxyethyl)phosphine (TCEP), or other reducing agents, and/or photolysis, as well as an ester bond can be cleaved for example by acidic or basic hydrolysis.
  • TCEP tris(2-carboxyethyl)phosphine
  • Locked Nucleic Acids As used herein, “locked nucleic acids” or “LNAs” (also known as 2’-O,4’-C-methylene-bridged nucleic acid (2’,4’- BNA)) are artificial nucleic acid derivatives that contain a methylene bridge connecting the 2’-O with the 4’-C position in the furanose ring. This enables them to form a strictly N-type conformation that offers high binding affinity against complementary RNA. LNA also presents enzyme resistance, similar to other nucleic acid derivatives. LNAs are used for various gene silencing techniques, such as antisense, short interfering RNA, blocking of microRNA, and triplex-forming oligonucleotides.
  • LNAs can be used, for example, in Splice Switching Oligonucleotides (SSOs) and LNA-based SSOs (LNA SSOs) are functional in vivo in mouse models. (See Shimo et. Al. Nucleic Acids Research, 42(12): 8174–8187 (2014), incorporated by reference herein in its entirety.) [0377] Modified: As used herein “modified” refers to a changed state or structure of a molecule of the disclosure. Molecules may be modified in many ways including chemically, structurally, and functionally. In some embodiments, the nucleic acid-based compositions of the present disclosure are modified by the introduction of non-natural nucleosides and/or nucleotides.
  • nucleic acid refers to a molecule comprised of one or more oligomeric polymers of at least 2 nucleoside monomers (nucleotide monomer when linked to another nucleoside. Each nucleoside monomer is further defined generally as comprising a sugar, nucleobase and backbone linker. Each of the sugar, nucleobase or backbone linker may be naturally occurring or synthetic.
  • the nucleosides or nucleotides can be ribonucleotides, deoxyribonucleotides, or both.
  • linkages may include any of the linkages known in the art including, for example, nucleic acids comprising 5' to 3' linkages.
  • the nucleotides may be naturally occurring or may be synthetically produced analogs that are capable of forming base-pair relationships with naturally occurring base pairs.
  • non-naturally occurring bases that are capable of forming base-pairing relationships include, but are not limited to, aza and deaza pyrimidine analogs, aza and deaza purine analogs, and other heterocyclic base analogs, wherein one or more of the carbon and nitrogen atoms of the pyrimidine rings have been substituted by heteroatoms, e.g., oxygen, sulfur, selenium, phosphorus, and the like.
  • patient refers to a subject who may seek or be in need of treatment, requires treatment, is receiving treatment, will receive treatment, or a subject who is under care by a trained professional for a particular disease or condition.
  • Peptide As used herein, “peptide” is less than or equal to 50 amino acids long, e.g., about 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50 amino acids long.
  • PNA Peptide nucleic acids
  • PNA Peptide nucleic acids
  • compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
  • pharmaceutically acceptable excipients refers any ingredient other than the compounds described herein (for example, a vehicle capable of suspending or dissolving the active compound) and having the properties of being substantially nontoxic and non-inflammatory in a patient.
  • Excipients may include, for example: antiadherents, antioxidants, binders, coatings, compression aids, disintegrants, dyes (colors), emollients, emulsifiers, fillers (diluents), film formers or coatings, flavors, fragrances, glidants (flow enhancers), lubricants, preservatives, printing inks, sorbents, suspensing or dispersing agents, sweeteners, and waters of hydration.
  • antiadherents antioxidants, binders, coatings, compression aids, disintegrants, dyes (colors), emollients, emulsifiers, fillers (diluents), film formers or coatings, flavors, fragrances, glidants (flow enhancers), lubricants, preservatives, printing inks, sorbents, suspensing or dispersing agents, sweeteners, and waters of hydration.
  • excipients include, but are not limited to: butylated hydroxytoluene (BHT), calcium carbonate, calcium phosphate (dibasic), calcium stearate, croscarmellose, crosslinked polyvinyl pyrrolidone, citric acid, crospovidone, cysteine, ethylcellulose, gelatin, hydroxypropyl cellulose, hydroxypropyl methylcellulose, lactose, magnesium stearate, maltitol, mannitol, methionine, methylcellulose, methyl paraben, microcrystalline cellulose, polyethylene glycol, polyvinyl pyrrolidone, povidone, pregelatinized starch, propyl paraben, retinyl palmitate, shellac, silicon dioxide, sodium carboxymethyl cellulose, sodium citrate, sodium starch glycolate, sorbitol, starch (corn), stearic acid, sucrose, talc, titanium dioxide, vitamin A, vitamin E, vitamin C,
  • compositions described herein also includes pharmaceutically acceptable salts of the compounds described herein.
  • pharmaceutically acceptable salts refers to derivatives of the disclosed compounds wherein the parent compound is modified by converting an existing acid or base moiety to its salt form (e.g., by reacting the free base group with a suitable organic acid).
  • examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like.
  • Representative acid addition salts include acetate, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptonate, hexanoate, hydrobromide, hydrochloride, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2- naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate,
  • alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like, as well as nontoxic ammonium, quaternary ammonium, and amine cations, including, but not limited to ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like.
  • the pharmaceutically acceptable salts of the present disclosure include the conventional non-toxic salts of the parent compound formed, for example, from non-toxic inorganic or organic acids.
  • the pharmaceutically acceptable salts of the present disclosure can be synthesized from the parent compound which contains a basic or acidic moiety by conventional chemical methods.
  • such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred. Lists of suitable salts are found in Remington’s Pharmaceutical Sciences, 17 th ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418, Pharmaceutical Salts: Properties, Selection, and Use, P.H. Stahl and C.G.
  • solvates means a compound of the disclosure wherein molecules of a suitable solvent are incorporated in the crystal lattice.
  • a suitable solvent is physiologically tolerable at the dosage administered.
  • solvates may be prepared by crystallization, recrystallization, or precipitation from a solution that includes organic solvents, water, or a mixture thereof.
  • solvents examples include ethanol, water (for example, mono-, di-, and tri-hydrates), N-methylpyrrolidinone (NMP), dimethyl sulfoxide (DMSO), N,N’- dimethylformamide (DMF), N,N’-dimethylacetamide (DMAC), 1,3-dimethyl-2- imidazolidinone (DMEU), 1,3-dimethyl-3,4,5,6-tetrahydro-2-(1H)-pyrimidinone (DMPU), acetonitrile (ACN), propylene glycol, ethyl acetate, benzyl alcohol, 2-pyrrolidone, benzyl benzoate, and the like.
  • NMP N-methylpyrrolidinone
  • DMSO dimethyl sulfoxide
  • DMF N,N’- dimethylformamide
  • DMAC N,N’-dimethylacetamide
  • DMEU 1,3-dimethyl-2- imidazolidinone
  • DMPU 1,3-
  • a “pharmacologic effect” is a measurable biologic phenomenon in an organism or system which occurs after the organism or system has been contacted with or exposed to an exogenous agent. Pharmacologic effects may result in therapeutically effective outcomes such as the treatment, improvement of one or more symptoms, diagnosis, prevention, and delay of onset of disease, disorder, condition or infection. Measurement of such biologic phenomena may be quantitative, qualitative or relative to another biologic phenomenon. Quantitative measurements may be statistically significant. Qualitative measurements may be by degree or kind and may be at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more different.
  • nucleic acid-based compositions of the present disclosure can comprise exogenous agents.
  • Examples of pharmacologic effects include, but are not limited to, alteration in cell count such as an increase or decrease in neutrophils, reticulocytes, granulocytes, erythrocytes (red blood cells), megakaryocytes, platelets, monocytes, connective tissue macrophages, epidermal langerhans cells, osteoclasts, dendritic cells, microglial cells, neutrophils, eosinophils, basophils, mast cells, helper T cells, suppressor T cells, cytotoxic T cells, natural killer T cells, B cells, natural killer cells, or reticulocytes.
  • alteration in cell count such as an increase or decrease in neutrophils, reticulocytes, granulocytes, erythrocytes (red blood cells), megakaryocytes, platelets, monocytes, connective tissue macrophages, epidermal langerhans cells, osteoclasts, dendritic cells, microglial cells, neutrophils, eosinophils, bas
  • Pharmacologic effects also include alterations in blood chemistry, pH, hemoglobin, hematocrit, changes in levels of enzymes such as, but not limited to, liver enzymes AST and ALT, changes in lipid profiles, electrolytes, metabolic markers, hormones or other marker or profile known to those of skill in the art.
  • Physicochemical means of or relating to a physical and/or chemical property.
  • the term “preventing” refers to partially or completely delaying onset of an infection, disease, disorder and/or condition; partially or completely delaying onset of one or more symptoms, features, or clinical manifestations of a particular infection, disease, disorder, and/or condition; partially or completely delaying onset of one or more symptoms, features, or manifestations of a particular infection, disease, disorder, and/or condition; partially or completely delaying progression from an infection, a particular disease, disorder and/or condition; and/or decreasing the risk of developing pathology associated with the infection, the disease, disorder, and/or condition.
  • Prognosing As used herein, the term “prognosing” means a statement or claim that a particular biologic event will, or is very likely to, occur in the future.
  • Progression As used herein, the term “progression” or “cancer progression” means the advancement or worsening of or toward a disease or condition.
  • Protein A "protein” means a polymer of amino acid residues linked together by peptide bonds. The term, as used herein, refers to proteins, polypeptides, and peptides of any size, structure, or function. Typically, however, a protein will be at least 50 amino acids long. In some instances, the protein encoded is smaller than about 50 amino acids.
  • polypeptide is termed a peptide. If the protein is a short peptide, it will be at least about 10 amino acid residues long.
  • a protein may be naturally occurring, recombinant, or synthetic, or any combination of these.
  • a protein may also comprise a fragment of a naturally occurring protein or peptide.
  • a protein may be a single molecule or may be a multi-molecular complex.
  • the term protein may also apply to amino acid polymers in which one or more amino acid residues are an artificial chemical analogue of a corresponding naturally occurring amino acid.
  • Protein expression refers to the process by which a nucleic acid sequence undergoes translation such that detectable levels of the amino acid sequence or protein are expressed.
  • sample As used herein, “purify,” “purified,” “purification” means to make substantially pure or clear from unwanted components, material defilement, admixture or imperfection.
  • sample As used herein, the term “sample” or “biological sample” refers to a subset of its tissues, cells or component parts (e.g. body fluids, including but not limited to blood, mucus, lymphatic fluid, synovial fluid, cerebrospinal fluid, saliva, amniotic fluid, amniotic cord blood, urine, vaginal fluid and semen).
  • body fluids including but not limited to blood, mucus, lymphatic fluid, synovial fluid, cerebrospinal fluid, saliva, amniotic fluid, amniotic cord blood, urine, vaginal fluid and semen).
  • a sample further may include a homogenate, lysate or extract prepared from a whole organism or a subset of its tissues, cells or component parts, or a fraction or portion thereof, including but not limited to, for example, plasma, serum, spinal fluid, lymph fluid, the external sections of the skin, respiratory, intestinal, and genitourinary tracts, tears, saliva, milk, blood cells, tumors, organs.
  • a sample further refers to a medium, such as a nutrient broth or gel, which may contain cellular components, such as proteins or nucleic acid molecule.
  • Single unit dose is a dose of any therapeutic administered in one dose/at one time/single route/single point of contact, i.e., single administration event.
  • Similarity refers to the overall relatedness between polymeric molecules, e.g. between polynucleotide molecules (e.g. DNA molecules and/or RNA molecules) and/or between polypeptide molecules. Calculation of percent similarity of polymeric molecules to one another can be performed in the same manner as a calculation of percent identity, except that calculation of percent similarity takes into account conservative substitutions as is understood in the art.
  • small interfering RNA As used herein “small interfering RNA” or “siRNA” mean a double-stranded RNA typically 20-25 nucleotides long involved in the RNA interference (RNAi) pathway and interfering with or inhibiting the expression of a specific gene. The gene is the target gene of the siRNA. A siRNA is usually about 21 nucleotides long, with 3' overhangs (e.g., 2 nucleotides) at each end of the two strands. A siRNA inhibits target gene expression by binding to and promoting the cleavage of one or more RNA transcripts of the target gene at specific sequences.
  • RNAi RNA interference pathway
  • Splice-Switching Oligonucleotides As used herein, “Splice-switching oligonucleotides” or “SSOs” are oligonucleotides that modulate pre-mRNA splicing, can repair defective RNA, and restore the production of essential proteins. They can also generate novel proteins with desirable properties and regulate the presence of disease-related splice variant proteins. The latter outcome may be achieved by modulating alternative splicing of pre-mRNA.
  • SSOs may block RNA sequences that are essential for splicing and prevent the interaction of splicing factors, such as RNA- binding proteins, small nuclear RNAs and other components of the spliceosome, with the pre- mRNA.
  • Split dose As used herein, a “split dose” is the division of single unit dose or total daily dose into two or more doses.
  • Stable As used herein “stable” refers to a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and in some embodiments, capable of formulation into an efficacious therapeutic agent.
  • Stabilized As used herein, the term “stabilize”, “stabilized,” “stabilized region” means to make or become stable.
  • Subject As used herein, the term “subject” or “patient” refers to any organism to which a composition in accordance with the disclosure may be administered, e.g., for experimental, diagnostic, prophylactic, and/or therapeutic purposes. Typical subjects include animals (e.g., mammals such as mice, rats, rabbits, non-human primates, and humans) and/or plants.
  • Substantially As used herein, the term “substantially” refers to the qualitative condition of exhibiting total or near-total extent or degree of a characteristic or property of interest.
  • Susceptible to An individual who is “susceptible to” a disease, disorder, and/or condition has not been diagnosed with and/or may not exhibit symptoms of the disease, disorder, and/or condition but harbors a propensity to develop a disease or its symptoms.
  • an individual who is susceptible to a disease, disorder, and/or condition may be characterized by one or more of the following: (1) a genetic mutation associated with development of the disease, disorder, and/or condition; (2) a genetic polymorphism associated with development of the disease, disorder, and/or condition; (3) increased and/or decreased expression and/or activity of a protein and/or nucleic acid associated with the disease, disorder, and/or condition; (4) habits and/or lifestyles associated with development of the disease, disorder, and/or condition; (5) a family history of the disease, disorder, and/or condition; and (6) exposure to and/or infection with a microbe associated with development of the disease, disorder, and/or condition.
  • an individual who is susceptible to a disease, disorder, and/or condition will develop the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition will not develop the disease, disorder, and/or condition.
  • Sustained release As used herein, the term “sustained release” refers to a pharmaceutical composition or compound release profile that conforms to a release rate over a specific period of time.
  • Synthetic The term “synthetic” means produced, prepared, and/or manufactured by the hand of man. Synthesis of polynucleotides or polypeptides or other molecules of the present disclosure may be chemical or enzymatic.
  • Targeted cells refers to any one or more cells of interest.
  • the cells may be found in vitro, in vivo, in situ or in the tissue or organ of an organism.
  • the organism may be an animal, in some embodiments, a mammal, or a human and most.
  • the targeted cells are cells of the kidney.
  • the targeted cells can be a subpopulation of cells of the kidney.
  • the targeted cells are kidney tubule cells.
  • Therapeutic Agent refers to any agent that, when administered to a subject, has a therapeutic, diagnostic, and/or prophylactic effect and/or elicits a desired biological and/or pharmacological effect.
  • therapeutically effective amount means an amount of an agent to be delivered (e.g., nucleic acid, drug, therapeutic agent, diagnostic agent, prophylactic agent, etc.) that is sufficient, when administered to a subject suffering from or susceptible to an infection, disease, disorder, and/or condition, to treat, improve symptoms of, diagnose, prevent, and/or delay the onset of the infection, disease, disorder, and/or condition.
  • an agent to be delivered e.g., nucleic acid, drug, therapeutic agent, diagnostic agent, prophylactic agent, etc.
  • Therapeutically effective outcome means an outcome that is sufficient in a subject suffering from or susceptible to an infection, disease, disorder, and/or condition, to treat, improve symptoms of, diagnose, prevent, and/or delay the onset of the infection, disease, disorder, and/or condition.
  • Total daily dose As used herein, a “total daily dose” is an amount given or prescribed in 24 hour period. It may be administered as a single unit dose.
  • Treating refers to partially or completely alleviating, ameliorating, improving, relieving, delaying onset of, inhibiting progression of, reducing severity of, and/or reducing incidence of one or more symptoms or features of a particular infection, disease, disorder, and/or condition.
  • “treating” cancer may refer to inhibiting survival, growth, and/or spread of a tumor.
  • Treatment may be administered to a subject who does not exhibit signs of a disease, disorder, and/or condition and/or to a subject who exhibits only early signs of a disease, disorder, and/or condition for the purpose of decreasing the risk of developing pathology associated with the disease, disorder, and/or condition.
  • a method of treating when applied to, for example, cancer refers to a procedure or course of action that is designed to reduce, eliminate or prevent the number of cancer cells in an individual, or to alleviate the symptoms of a cancer.
  • a method of treating does not necessarily mean that the cancer cells or other disorder will, in fact, be completely eliminated, that the number of cells or disorder will, in fact, be reduced, or that the symptoms of a cancer or other disorder will, in fact, be alleviated.
  • a method of treating cancer will be performed even with a low likelihood of success, but which, given the medical history and estimated survival expectancy of an individual, is nevertheless deemed an overall beneficial course of action.
  • Unmodified refers to any substance, compound or molecule prior to being changed in any way. Unmodified may, but does not always, refer to the wild type or native form of a biomolecule. Molecules may undergo a series of modifications whereby each modified molecule may serve as the “unmodified” starting molecule for a subsequent modification.
  • Upregulation refers to an increase in the level of expression of the gene, an increase in the level of the polypeptide(s) encoded by the gene, the level bioactive gene product, or level of the RNA transcript(s) transcribed from the template strand of the gene as compared to the levels observed in the absence of the nucleic acid-based composition(s) described herein.
  • the nucleic acid-based compositions of the present disclosure may have a direct upregulating effect on the expression of the target gene.
  • upregulation depending on the context, can also refer to the effect of a trans-acting activator of expression. VII.
  • compositions of the disclosure can be excluded from any one or more claims, for any reason, whether or not related to the existence of prior art.
  • All cited sources for example, references, publications, databases, database entries and art cited herein are incorporated into this application by reference even if not expressly stated in the citation. In case of conflicting statements of a cited source and the instant application, the statement in the instant application shall control.
  • the disclosure is further illustrated by the following non-limiting examples. It is to be understood that the foregoing description and following examples are intended to illustrate and not limit the scope of the present disclosure, which is defined by the scope of the appended claims.
  • Split site analysis identified chr15:50570778- 50570829 of the 3’ UTR and chr15:50570981-50570999 of the intron between exon 9 and exon 8 to be enriched for potential splicing motifs.
  • the final target sequence used for oligonucleotide tiling design was chr15:50570778-50570999 (this sequence is provided as SEQ ID NO: 5).
  • Figure 1 shows the exon-intron structure of GABPB1L and GABPB1S (not to scale).
  • the inset shows exon 9 of GABPB1L as well as the flanking intron and UTR sequences.
  • Putative splice motifs identified by computational analysis are represented by (*).
  • the tiling window is defined by the vertical dotted lines, and candidate oligonucleotide sequences are represented by the horizontal black lines. Oligonucleotide designs that passed additional filtering criteria advanced to transfections in LN229-P2A-NLuc cells to test their ability to silence TERT expression.
  • Each oligo candidate had Phosphorothioate backbones and Locked-Nucleic Acids (LNA) incorporated into their design to generate an antisense LNA oligo candidate (ASO) or a splice-switching LNA oligonucleotide candidate (SSO).
  • LNA Locked-Nucleic Acids
  • siRNA Design The target sequence for siRNA design included exon 9 of GABPB1L as well as the same portion of the 3’UTR that was scanned for ASOs (chr15:50570778-50570829). In addition, 11 base pairs of the flanking exon 8 sequence was included so that putative siRNAs spanning the exon 8-9 junction could be identified. The full cDNA sequence that was used to scan for siRNA is represented by SEQ ID NO: 6. [0434] This cDNA sequence was scanned for putative functional siRNA sequence using the selection criteria described in Reynolds et al., Nat. Biotechnol.
  • Pattern 1 was AA(N19)TT (SEQ ID NO: 10) and Pattern 2 was NA(N21) (SEQ ID NO: 11).
  • N any nucleotide. This yielded 81 siRNA candidate sequences.
  • siGenome SMART pools were ordered as control siRNAs.
  • siGABPB1 M-013083-01-0005
  • siTERT M-003547-02-0005
  • non-targeting siCTRL D-001206-13-05
  • ASOs, SSOs, and ASO duplexes were all ordered from Integrated DNA Technologies (website: www.idtdna.com). ASOs and SSOs were ordered at 100nmol scale with standard desalting. siRNA sense and antisense RNA strands were ordered at 100nmol scale and underwent hybridization and concentration normalization prior to delivery.
  • siRNA Primary Screen [0437] All candidate siRNAs were initially transfected into a glioblastoma (GBM) reporter cell line (LN229-P2A-NLuc) expressing NanoLuc Luciferase endogenously through a P2A fusion with the endogenous TERT gene.
  • GBM glioblastoma
  • LN229-P2A-NLuc was prepared as follows: [0438] The NanoLuc Luciferase protein was generated as an in-frame fusion with the endogenous TERT gene on the same allele containing the C228T TERT promoter mutation. A homologous recombination knock-in approach was used.
  • TERT- P2A-NLuc-TERT The final 300bp of the TERT coding sequence, followed by a P2A self-cleaving peptide sequence (derived from Porcine teschovirus-12A), followed by NanoLuc Luciferase, followed by the first 300bp of the TERT 3’ UTR were cloned into the pUC57 vector in sequence (plasmid referred to as TERT- P2A-NLuc-TERT).
  • CRISPR guide RNAs designed to cut at the TERT coding sequence- 3’UTR junction were cloned into the px459 vector (Genscript, Piscataway, NJ, Cat No.
  • TERT-g1-px459 plasmid referred to as TERT-g1-px459
  • TERT-g1-px459 plasmid referred to as TERT-g1-px459
  • TERT-g1-px459 and TERT-P2A-NLuc-TERT plasmids were co-transfected using the lipofectamine 3000 system (ThermoFisher, Waltham, MA, Cat No. L3000001). 48 hours post transfection, 1 ⁇ g/ml of Puromycin was added to the cells to select for cells that had successfully taken up the TERT-g1-px459 plasmid.
  • the cells were propagated for 6 days in the presence of puromycin. On day 6, the cells were trypsinized. Half of the cells were re-seeded in 96 well plates at a density of 20 cells/well while the other half were used to collect genomic DNA. Junction PCR was performed on the genomic DNA to detect the presence of cells containing the correct knock- in at the TERT locus. After two weeks of expansion, NanoLuc Luciferase was measured in each well of the 96-well plate and the wells with a positive signal were advanced to single cell cloning. [0442] Single-cell clones containing the correct integration were derived and frozen down for future use (termed LN229-P2A-Nluc hereafter).
  • ASO Primary Screen All candidate ASOs were initially transfected into the LN229 GBM cell line, containing the C228T TERTp mutation. Cells were seeded at 4,000 cells/ well in 96-well plates 24 hours prior to transfection. The following day, ASOs were transfected at 25nM. All transfections were performed with Dharmafect1 (Dharmacon, Lafayette, CO, T-2001-01), with 0.15uL of Dharmafect 1 being used for each well.
  • FIG. 2 provides a set of charts showing the expression fold change, nano luciferase and fluorescence (590 nm) for the samples at 72 hours.
  • LN229 GBM cells (TERTp mutant) and LN229-P2A-NLuc cells were transfected with 50nM of candidate siRNA.
  • 72 hours post drug treatment cell lysates were harvested and TERT mRNA was measured via RT-qPCR (ThermoFisher, Waltham, MA, Cat. No. A25600).
  • TSiR-1, TSiR-37, TSiR-26, and TSiR-34 were selected as hits for displaying a reduction in NanoLuc Luciferase activity at either 48 or 72 hours.
  • TSiR-6 was randomly selected from the candidates that had no effect on reporter activity.
  • siRNA hit Confirmation 24-hours prior to transfection, 4,000 cells/well of LN229 GBM cells were seeded into two 96-well plates. A matching plate of LN229-P2A-NLuc cells were also seeded. Triplicate wells were used for each condition, and each siRNA was transfected as described above. 72-hours post transfection, RNA was harvested from plate 1, plate 2 was used to measure cell-viability (ThermoFisher PrestoBlue: A13261), and the LN229-P2A-NLuc plate was used to measure NanoLuc Luciferase.
  • RT-qPCR analysis was performed on all collected RNA lysates to determine relative expression changes in GABPB1L and TERT mRNA (GUSB primers were used as an internal control).
  • siRNA Cell Panel Analysis [0450] To test if any of the siRNA hits could reduce TERT expression selectively in cancer cells harboring TERTp mutations, we performed additional transfections in a panel of two TERTp-mutant and two TERTp wild-type cell lines. LN18 is a GBM cell line that expresses TERT from the wild-type promoter, and 293T cells are TERTp wild-type cells derived from embryonic kidney cells.
  • LN229 is GBM cell line and HepG2 is a Hepatocellular carcinoma cell line, both of which are positive for the C228T TERTp mutation.
  • Each cell line underwent a transfection optimization to determine the ideal amount of Dharmafect 1 transfection reagent to achieve maximal GABPB1L knockdown while minimizing cell toxicity. It was found that 0.2 ⁇ L/well was ideal for LN229 and LN18, 0.3 ⁇ L/well was optimal for 293T, and 0.4 ⁇ L/well was optimal for HepG2 cells.
  • TSiR-1, TSiR-37, TSiR-26, and TSiR-34 were transfected at 50nM into the four cell lines in 96-well plates.
  • siCTRL spiked non-targeting
  • siGABPB1 siGABPB1 were used as negative and positive control siRNAs respectively.
  • 72 hours post transfection the cells were harvested for RNA and changes in GABPB1L and TERT mRNA ( Figure 3 and Figure 4, respectively) were determined by RT-qPCR analysis.
  • Both TSiR-1 and TSiR-34 achieved a greater TERT reduction in the two TERTp mutant lines when compared to the TERTp wild- type cell lines.
  • this effect was particularly pronounced as the knockdown efficiency of GABPB1L was greatest in LN18 (WT) and weakest in HepG2 (Mut).
  • TAO-21, TAO-22, TAO-37, and TAO-40 were selected from the primary screen as hits for displaying greater than 60% reduction in TERT mRNA at 72 hours.
  • LN18 is a GBM cell line that expresses TERT from the wild- type promoter, and 293T cells are TERTp wild-type cells derived from embryonic kidney cells.
  • LN229 is GBM cell line and HepG2 is a Hepatocellular carcinoma cell line, both of which are positive for the C228T TERTp mutation.
  • the four ASO hits were transfected at 25nM into the four cell lines in 96-well plates.
  • a scrambled non-targeting LNA-ASO (Scr) was used as a negative control.
  • 72 hours post transfection the cells were harvested for RNA and changes in TERT mRNA were determined by RT-qPCR analysis ( Figure 5).
  • TAO-21, TAO-22, and TAO-37 achieved a greater TERT reduction in the two TERTp mutant lines when compared to the TERTp wild- type cell lines.
  • GABPB1-hybridizing oligonucleotides All possible 20-mer GABPB1-hybridizing oligonucleotides (MOE, 5-10-5) were designed to target GABPB1L or total GABPB1 (both the GABPB1L and GABPB1S isoforms).
  • the GABPB1-hybridizing oligonucleotides that were designed to target GABPB1L are shown in Table 5 and the GABPB1-hybridizing oligonucleotides that were designed to target total GABPB1 are shown in Table 6.
  • Screen of 150 ASOs [0456] Based on the bioinformatics analysis of the oligonucleotides in Table 5 and Table 6, 150 ASOs (Table 11) were selected for screening.
  • the candidate ASOs were transfected into Hep3B cells which were seeded at 15,000 cells/well in 96-well plates. The following day, ASOs were transfected at 20 nM or 2 nM. 24 hours post transfection, the cells were harvested and GABPB1L was measured with a bDNA assay. The mean relative GABPB1L mRNA level (“Relative mRNA Expression”) for the 20 nM and 2 nM doses for each ASO tested is shown in Table 11 as well as the cell viability for the 20 nM dose. Table 11. Mean Relative GABPB1L mRNA
  • 50 ASOs were selected for additional studies. Screen of 50 ASOs [0458] 50 ASOs were selected from the screen of the 150 ASOs. The candidate ASOs were transfected into Hep3B cells which were seeded at 15,000 cells/well in 96-well plates. The following day, ASOs were transfected at 20 nM or 5 nM. 72 hours post transfection, the cells were harvested and GABPB1L and TERT mRNA was measured by RT-qPCR. The knockdown percentage for GABPB1L and TERT are in shown in Table 12 as well as the cell viability for the 5 nM dose. Table 12. GABPB1L and TERT mRNA Expression
  • TAO-A2641.1, TAO-A4902.1, TAO-A5551.1, TAO-A5552.1, TAO-A6451.1, TAO-A6453.1, TAO-A6529.1, TAO-A6577.1, TAO-A7186.2, TAO-A7210.1, TAO- A7215.1, TAO-A7441.1, TAO-A7797.1, TAO-A8210.1, TAO-A10683.1, TAO-A11082.1, and TAO-A11083.1 (collectively referred to as “17 ASOs”) were selected from the screen of the 50 ASOs described in Table 12 for further screening and to design the additional 16-mer, 18-mer and 20-mer ASOs shown in Table 8.
  • the 17 ASOs and the 63 ASOs from Table 12, (a total of 80 ASOs) were transfected into Hep3B cells which were seeded at 15,000 cells/well in 96-well plates. The following day, ASOs were transfected at 20 nM or 5 nM. 24 hours post transfection, the cells were harvested and mRNA levels were measured by bDNA assay for hsGABPB1L, hsCDKN1A (hsGABPB1L silencing should lead to an upregulation of hsCDKN1A) and hsGAPDH. The mean value (MV) of the remaining mRNA levels GAPDH percentage (GAPDH %) for each ASO is shown in Table 13. Table 13.
  • a series of screens were performed to identify oligonucleotides that decrease GABPB1L (the beta1L tetramer-forming isoform of GABP) and/or TERT mRNA expression in vitro.
  • TAO-95.1 to TAO-122.1 U87, Huh7, LN229 and 293T Cells
  • TAO-95.1 to TAO-122.1 were synthesized and transfected into U87 and Huh7 cells that are both positive for the C228T mutation.
  • TAO-95.1, TAO-97.1, TAO-104.1, TAO-106.1, TAO-115.1, TAO-116.1, TAO-118.1 and TAO-120.1 were selected for additional studies in LN229 (GBM cell line) and 293T (TERTp wild-type cells derived from embryonic kidney cells) cells.
  • the average cell viability and the average TERT mRNA fold change for the LN229 and the 293T cells after 72 hours is provided in Table 15.
  • a comparison of the ASOs across the 4 cell lines (Huh7, U87, LN229 and 293T) is shown in Table 16.
  • TAO-37.1 was used as a positive control.
  • Table 15 Cell Viability and TERT mRNA Fold Change in LN229 and 293T Cells Table 16. Cell Viability and TERT mRNA Fold Change Across Huh7, U87, LN229 and 293T Lines TAO-123.1 to TAO-139.1: Huh7 and LN229 Cells [0464] TAO-123.1 to TAO-139.1 where were synthesized and transfected into Huh7 and LN229 cells that are both positive for the C228T mutation.
  • TAO-123.1 and TAO-125.1 are locked nucleic acids (LNAs) that have a backbone that includes phosphorothioate (Ps) and phosphodiester (Po) bonds
  • TAO-124.1 is an LNA that has a backbone that includes Ps bonds
  • TAO-126.1 to TAO-139.1 include 2’-O-(2-Methoxyethyl)-modified nucleotides (2-MOE) and backbones that include Ps and Po bonds.
  • An initial screen using a PrestoBlue® assay was performed to determine cell viability and qPCR was used to measure the levels of GABPB1L and TERT mRNA.
  • the average cell viability, and the average GABPB1L mRNA and TERT mRNA fold change at 48 hours is provided in Table 17.
  • a scrambled non-targeting LNA-ASO (ScrASO) was used as a negative control, and TAO-115.1 and TAO-95.1 were used as a positive control.
  • Table 17. Cell Viability, TERT mRNA and GABPB1L Fold Change in Huh7 and LN229 Cells (48 Hours)
  • TAO- 123.1, TAO-125.1, TAO-130.1, TAO-131.1, TAO-132.1, TAO-135.1, TAO-137.1 and TAO- 139.1 were selected for a follow-up time course study.
  • the average cell viability, and the average GABPB1L mRNA and TERT mRNA fold change at 96 hours is provided in Table 18.
  • Table 19 Provided in Table 19 is a summary of the time course data for the LN229 cells.
  • a scrambled non-targeting LNA-ASO (ScrASO) was used as a negative control along with an untreated sample, and TAO-115.1 and TAO-95.1 were used as a positive control.
  • Table 20 shows the average cell viability where DMSO was used as the vehicle only control and the results are shown as percent viability from vehicle. Shown in Table 21 are the average GABPB1L mRNA fold change as compared to the vehicle only and Table 22 are the average TERT mRNA fold change as compared to the vehicle only. For TAO-37.1 and TAO-115.1, the mRNA expression fold change for both GABPB1L and TERT was lowest for the higher dose and was highest at the lowest dose. For TAO-98.1 this trend was only seen for TERT expression. Table 20. Cell Viability for TAO-37.1, TAO-98.1, TAO-115.1
  • Example 6 In Vivo Study [0468] Various in vivo studies were conducted to evaluate a subset of the ASOs described herein. CED Delivery of ASO [0469] Three ASOs (TAO-115.1, TAO-37.1 and TAO-98.1) were selected for in vivo studies in mice. Mice were inoculated on day 1 with tumor and then administered 5 uL of TAO-115.1, TAO-37.1 and TAO-98.1 or a control of scrambled non-targeting LNA-ASO (ScrASO) by convection enhanced delivery (CED). On day 6 the tumors were harvested and analyzed by RT-qPCR.
  • ScrASO scrambled non-targeting LNA-ASO
  • TAO-115.1 The average and median relative expression (Fold Change to ScrASO) of GABPB1 and TERT are shown in Table 23 and Table 24 respectfully.
  • mice were inoculated with tumor, BLI was measured at day 7 and day 12, and then on day 12 the mice were administered by intratumoral (ITu) delivery a single dose of 300 ug ScrASO, 300 ug of TAO-115 or 500 ug of TAO-115.
  • mice were inoculated with tumor, BLI was measured at day 7 and day 12, and then on day 12, day 14 and day 16 the mice were administered by intratumoral (ITu) delivery a dose of 300 ug ScrASO, 200 ug of TAO-115.1 or 300 ug of TAO-115.1.
  • MV mean value of cell viability
  • GABPB1L mRNA The mean value (MV) of cell viability, GABPB1L mRNA, and TERT mRNA data are shown in Table 26.
  • Table 26. Cell Viability and GABPB1 and TERT mRNA % Change * The negative value means the mRNA level was reduced and the positive value means the mRNA level was increased.

Abstract

Provided herein are compositions and methods for treating cancer by down-regulating telomerase. In particular, the present disclosure provides nucleic acids that inhibit GABPB1 expression. In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target GABPB1L, GABPB1S or total GABPB1 mRNA and reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations.

Description

NUCLEIC ACIDS FOR INHIBITING TERT EXPRESSION CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims priority to US Provisional Application No. 63/071,795, filed August 28, 2020, entitled NUCLEIC ACIDS FOR INHIBITING TERT EXPRESSION, the contents of which are incorporated herein by reference in its entirety. SEQUENCE LISTING [0002] The present application is being filed along with a Sequence Listing in electronic format. The Sequence Listing file, entitled 2192_1001PCT_SL.txt, was created on August 26, 2021, and is 5,873,265 bytes in size. The information in electronic format of the Sequence Listing is incorporated herein by reference in its entirety. FIELD OF DISCLOSURE [0003] The disclosure relates to compositions and methods for treating cancer by downregulating telomerase. In particular, the disclosure provides nucleic acids that inhibit GABPB1 expression (e.g., partial GABPB1 expression or total GABPB1 expression). In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target exon 9 of GABPB1L mRNA and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations. In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target the intron between exon 8 and exon 9 of GABPB1L mRNA and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations. In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target total GABPB1 by targeting both GABPB1S and GABPB1L isoforms of the GABPB1 gene, and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations. BACKGROUND [0004] Telomerase expression is a hallmark of tumorigenesis. Due to its fundamental nature in driving tumorigenesis, many attempts have been made to inhibit telomerase as a cancer therapeutic strategy, but thus far none have become a standard of care. One promising approach was the oligonucleotide therapy GRN163L from Geron, Inc. By hybridizing and inhibiting the RNA template of telomerase, GRN163L reduced tumor growth in preclinical models of breast cancer, glioblastoma (GBM), pancreatic, and liver cancer. This preclinical success has not translated to the clinic however, as trials in breast, lung, and pediatric CNS cancers were discontinued. In each case, a high frequency of grade III/IV hematopoietic toxicities were observed. This was thought to result from inhibiting telomerase activity in healthy hematopoietic stem cells. As a result, trials with GRN163L have been restricted to myeloproliferative diseases. Therefore, there is currently a large unmet need to effectively inhibit telomerase activity selectively in cancer cells. SUMMARY OF DISCLOSURE [0005] The present disclosure provides compositions and methods for treating cancer by downregulating telomerase. In particular, the disclosure provides nucleic acids that inhibit GABPB1 expression (e.g., partial GABPB1 expression or total GABPB1 expression). In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target exon 9 of GABPB1L mRNA and reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations. In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target the intron between exon 8 and exon 9 of GABPB1L mRNA and reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations. In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target total GABPB1 by targeting both GABPB1S and GABPB1L isoforms of the GABPB1 gene, and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations. [0006] In some embodiments, the present disclosure provides an oligonucleotide capable of hybridizing to a target sequence. The target sequence may comprise exon 9 or the intron between exon 8 and 9 of a GABPB1 gene (DNA) or gene product (RNA) and the target sequence may be, but not limited to, SEQ ID NOs: 308-346, and 12424-24399. The oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 50% of the oligonucleotide sequence. The oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 75% of the oligonucleotide sequence. The oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 85% of the oligonucleotide sequence. The oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 90% of the oligonucleotide sequence. The oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over at least 99% of the oligonucleotide sequence. The oligonucleotide may be a sequence of 8-20 nucleotides in length and hybridization may occur over 100% of the oligonucleotide sequence. The oligonucleotide may include one or more modified nucleobases, sugars and/or linkers such as, but not limited to, LNAs, phosphorothioate (Ps) linkages, phosphodiester (Po) linkages, 5- methylcytosine modified nucleotides, and 2’MOE modified nucleotides. [0007] In some embodiments, the present disclosure provides a double-stranded small inhibitory RNA (siRNA) comprising the sequence of any one of SEQ ID NO: 386-424. [0008] In some embodiments, the present disclosure provides a method of reducing the activity of GA binding protein transcription factor subunit beta 1 (GABPB1) in a cell, comprising exposing the cell to any of the oligonucleotides, siRNA, ASOs, and pharmaceutical compositions thereof which are disclosed herein. In some aspects, the activity of GABPB1L isoform, GABPB1S isoform or total GABPB1 (both GABPB1L and GABPB1S isoforms) may be reduced. [0009] In some embodiments, the present disclosure provides a method of reducing the expression of telomerase reverse transcriptase (TERT) in a cell having a mutant TERT promoter with one or more somatic mutations, comprising exposing the cell to any of the oligonucleotides, siRNA, ASOs, and pharmaceutical compositions thereof which are disclosed herein. [0010] In some embodiments, the present disclosure provides a pharmaceutical composition, comprising any of the oligonucleotides, siRNA or ASOs and a pharmaceutically-acceptable carrier. The present disclosure provides a medicament, comprising any of the oligonucleotides, siRNA or ASOs and a pharmaceutically-acceptable carrier. [0011] In some embodiments, the present disclosure provides a method of treating cancer in a subject comprising administering the pharmaceutical compositions described herein to the subject. [0012] In some embodiments, the present disclosure provides a use of any of the siRNA disclosed herein for the manufacture of a pharmaceutical composition for treating cancer. [0013] In some embodiments, the present disclosure provides any of the siRNA disclosed herein for use in treating cancer. [0014] In some embodiments, the present disclosure provides an antisense oligonucleotide (ASO) which may include any of SEQ ID NO: 19-111, 430-474, 12190-12339, and 12361- 12423. As a nonlimiting example, the ASO may comprise SEQ ID NO: 12335, 12338, 12363, 12372, 12376, 12379, 12382, 12383, 12384, 12389, 12393, 12394, 12395, 12413, 12421, and 12422. [0015] In some embodiments, the present disclosure provides a method of reducing the activity of GA binding protein transcription factor beta 1 (GABPB1) in a cell, comprising exposing the cell to any of the ASOs described herein. In some aspects, the activity of GABPB1L isoform, GABPB1S isoform or total GABPB1 (both GABPB1L and GABPB1S isoforms) may be reduced. [0016] In some embodiments, the present disclosure provides a method of reducing the expression of telomerase reverse transcriptase (TERT) in a cell having a mutant TERT promoter with one or more somatic mutations, comprising exposing the cell to any of the ASOs described herein. [0017] In some embodiments, the present disclosure provides a pharmaceutical composition, comprising any of the ASOs described herein and a pharmaceutically- acceptable carrier. [0018] In some embodiments, the present disclosure provides a medicament, comprising any of the ASOs described herein and a pharmaceutically-acceptable carrier. [0019] In some embodiments, the present disclosure provides a method of treating a patient with cancer, comprising administering any of the pharmaceutical compositions described herein to the patient. [0020] In some embodiments, the present disclosure provides a use of any of the ASOs described herein for the manufacture of a pharmaceutical composition for treating cancer. [0021] In some embodiments, the present disclosure provides the ASOs described herein for use in treating cancer. DESCRIPTION OF THE DRAWINGS [0022] Figure 1 shows the exon-intron structure of GABPB1L and GABPB1S (not to scale). The inset shows exon 9 of GABPB1L as well as the flanking intron and UTR sequences. Putative splice motifs identified by computational analysis are represented by (*). The tiling window is defined by the vertical dotted lines, and candidate oligonucleotide sequences are represented by the horizontal black lines. Oligonucleotide designs that passed additional filtering criteria advanced to transfections in LN229-P2A-NLuc cells to test their ability to silence TERT expression. [0023] Figure 2 provides a set of charts showing the expression fold change, nano luciferase and fluorescence (590 nm) for the samples. LN229 GBM cells (TERTp mutant) and LN229-P2A-NLuc cells were transfected with 50nM of candidate siRNA. 72 hours post drug treatment, cell lysates were harvested and TERT mRNA was measured via RT-qPCR (ThermoFisher, Waltham, MA, Cat. No. A25600). Replicate plates were used to measure cell viability (ThermoFisher PrestoBlue: A13261), and the LN229-P2A-NLuc cell line was used to measure NanoLuc Luciferase. All treatment samples were normalized to the vehicle control of their respective cell type. Error bars represent a standard deviation of three replicates. [0024] Figure 3 shows GABPB1L Expression. TsiR-1, TsiR-37, TsiR-26, and TsiR-34 were transfected at 50nM into two TERTp mutant (LN229 and HepG2) and two TERTp wild-type (LN18 and 293T) cell lines in 96-well plates. siCTRL (scrambled non-targeting) and siGABPB1 were used as negative and positive control siRNAs respectively. 72 hours post transfection, the cells were harvested for RNA and changes in GABPB1L mRNA were determined by RT-qPCR analysis (ThermoFisher, Waltham, MA, Cat. No. A25600). All treatment samples were normalized to the siCTRL of their respective cell type. Error bars represent a standard error of the mean from 2 replicates. [0025] Figure 4 shows TERT expression. TsiR-1, TsiR-37, TsiR-26, and TsiR-34 were transfected at 50nM into two TERTp mutant (LN229 and HepG2) and two TERTp wild-type (LN18 and 293T) cell lines in 96-well plates. siCTRL (scrambled non-targeting) and siGABPB1 were used as negative and positive control siRNAs respectively. 72 hours post transfection, the cells were harvested for RNA and changes in TERT mRNA were determined by RT-qPCR analysis (ThermoFisher, Waltham, MA, Cat. No. A25600). All treatment samples were normalized to the siCTRL of their respective cell type. Error bars represent a standard error of the mean from 2 replicates. [0026] Figure 5 shows TERT expression for specific ASOs in different cells lines. TAO- 21, TAO-22, TAO-37, and TAO-40 were transfected at 25nM into two TERTp mutant (LN229 and HepG2) and two TERTp wild-type (LN18 and 293T) cell lines in 96-well plates. A scrambled non-targeting LNA-ASO (Scr) was used as a negative control. 72 hours post transfection, the cells were harvested for RNA and changes in TERT mRNA were determined by RT-qPCR analysis (ThermoFisher, Waltham, MA, Cat. No. A25600). All treatment samples were normalized to the Scr of their respective cell type. Error bars represent a standard error of the mean from at least 2 replicate wells. [0027] The details of various embodiments of the disclosure are set forth in the description below. Other features, objects, and advantages of the disclosure will be apparent from the description, from the drawings, and from the claims. DETAILED DESCRIPTION [0028] The present disclosure provides compositions and methods for downregulating telomerase expression. Telomerase expression may be downregulated by modulating GA Binding Protein Transcription Factor Beta 1 (GABPB1) and/or or telomerase reverse transcriptase (TERT) gene expression and/or function. Modulation of GABPB1 and/or TERT gene expression, as used herein, includes decreasing translation of a functional gene product; i.e., decreasing the translation of, and hence the levels of, GABPB1 (total or partial GABPB1 by reducing the level of GABPB1S or GABPB1L) and/or TERT.
[0029] In some embodiments, the compositions and methods described herein involve administering GABPB1 -hybridizing oligonucleotides to subjects having a disease, disorder and/or condition such as, but not limited to, cancer. Telomerase expression is a hallmark of tumorigenesis and over 90% of human cancers aberrantly express the enzyme. Telomerase functions by elongating telomeres, the ‘TTAGGG’ DNA repeats at the end of chromosomes. The majority of normal tissues have no telomerase activity so that telomeres shorten with each successive round of cell division. Eventually, a critical telomere length is reached and cells enter replicative senescence or undergo apoptosis. Telomerase reverse transcriptase (TERT) is a catalytic subunit of telomerase which catalyzes the addition of nucleotides in a specific DNA sequence to the ends of a chromosome’s telomeres. This addition of repetitive DNA sequences prevents degradation of the chromosomal ends after multiple rounds of replication. Reactivation of telomerase reverse transcriptase (TERT) expression occurs in many human cancers and TERT reactivation is necessary to overcome replicative senescence (aging) and prevent apoptosis (cell death), both fundamental steps in the initiation of cancer. [0030] Thus, the GABPB1 -hybridizing oligonucleotides described herein can be administered to treat individual subjects having a disease, disorder and/or condition such as, but not limited to, cancer.
[0031] The compositions taught herein are nucleic acid-based compositions. As used herein, “nucleic acid-based” compositions are oligomeric polymers of at least 2 nucleoside monomers (nucleotide monomer when linked to another nucleoside. Each nucleoside monomer is further defined generally as comprising a sugar, nucleobase and backbone linker. Each of the sugar, nucleobase or backbone linker may be naturally occurring or synthetic. Certain nucleic acid-based compositions effect their outcomes through a hybridization mechanism. As used herein, a “hybridization mechanism” refers to the capacity of a first oligonucleotide to form a complementary or reverse complementary (e.g., antisense) association with a second oligonucleotide via standard or non-standard hydrogen bonding. The association may form along the entire length, a part, a region or intermittently along the first or second oligonucleotide.
[0032] Accordingly, the present disclosure embraces nucleic acid-based compositions which are oligonucleotides that hybridize to other oligonucleotides. These compositions and methods may comprise at least one antisense oligonucleotide (ASO) that downregulates the expression of the GABPB1 gene. In some embodiments, the nucleic acids are oligonucleotides or siRNAs that target GABPB1L mRNA and inhibit GABP binding to mutant TERT promoter sequences. The oligonucleotides or siRNAs may target exon 9 or the intron between exon 8 and exon 9 of GABPB1L mRNA and consequently reduce TERT mRNA expression in cancer cells harboring TERT promoter mutations. I. Telomerase reverse transcriptase (TERT) and GA Binding Protein Transcription Factor Beta 1 (GABPB1) [0033] The TERT gene (Genbank Acc. No. NC_000005.10; RefSeq ID: NM_198253.3 provided as SEQ ID NO: 1) encodes the catalytic subunit of telomerase and its transcriptional regulation is the rate-limiting step in telomerase activity. Certain mutations in the TERT promoter have been associated with cancer cells. The most commonly occurring TERT promoter mutations are located 124bp and 146bp upstream of the translation start site and referred to as C228T and C250T, respectively, based on their hg19 genomic coordinates (genome.ucsc.edu/cgi-bin/hgGateway?db=hg19). The mutations are typically heterozygous, occur in a mutually exclusive fashion, and both create an identical 11bp sequence. Cells harboring these mutations reactivate TERT gene expression and telomerase activity, thus becoming immortalized. (Horn et al., Science doi:10.1126/science.1230062 (2013); Huang et al., Science, doi:10.1126/science.1229259 (2013), each of which are incorporated by reference in their entirety). These hotspot mutations have now been identified in over 50 distinct cancer types. In some cancer indications, TERTp mutations are observed in a strikingly high percentage of patients including 83% of Glioblastoma (GBM), 78% of Oligodendroglioma, 66% of bladder cancers, and 44% of Hepatocellular Carcinoma. [0034] The present disclosure utilizes these two hotspot point mutations located at position 68 and 90 in the TERT promoter (TERTp, SEQ ID NO: 2). For these hotspot point mutations, one or both of the guanine residues which may be replaced with adenine are shown below:
Figure imgf000009_0001
provides the TERT promoter with G residues replaced by A for both of the hotspot mutations. [0035] The present disclosure recognizes that the GA binding protein transcription factor beta 1 (GABPB1) activates the mutant TERTp in cancer cells. GABP binds and regulates the mutant TERT promoter but not the wild-type promoter in the same cells. Furthermore, GABP binding to the mutant TERT promoter is a common mechanism of TERT activation across multiple cancer types including glioblastoma (GBM), melanoma, hepatocellular carcinoma, and bladder cancer. (Bell et al., Science 348:1036–1039 (2015), S. Borah et al., Science 347:1006–1010 (2015), Amen et al. bioRxiv (2020) doi: doi.org/10.1101/2020.04.25.061606, each of which are incorporated by reference herein in their entirety). Thus, the present disclosure provides therapies that inhibit this mutation-specific interaction in at least the more than 50 distinct cancers that have been observed to harbor TERTp mutations to date. Many of these TERTp mutant cancers, such as GBM, have a high unmet medical need. Furthermore, contemplated herein are methods for treating cancers having TERTp mutations that have not yet been associated with this etiology. [0036] GABP is a unique transcription factor within the E26 Transformation Specific (ETS) family because it must either form a heterodimer (GABPA+GABPB), or heterotetramer (two A/B dimers), in order to bind DNA and activate transcription. The GABPA gene encodes a single protein isoform (Genbank Accession: NC_000021.9) that binds to DNA but cannot activate transcription. In contrast, GABPB1 encodes multiple protein isoforms that have a transcriptional activation domain but cannot bind to DNA. Not every isoform of GABPB1 allows for tetramer formation. While the GABPB1L isoform (Genbank Accession: NC_000015.10; chr15:50570778-50570999; the Hg19 coordinates for exon 9 of GABPB1L are chr15:50570829-50570981 and SEQ ID NO: 4 provides the GABPB1L Exon 9 coding sequence; GABPB1L Exon 9 coding sequence, splice sites, UTR provided as SEQ ID NO: 5) can form a tetramer via a Leucine Zipper Domain (LZD) at the C-terminus, the GABPB1S isoform lacks this domain and cannot dimerize to another GABPB1 subunit. [0037] In certain embodiments, at least one of the nucleotide substitutions in a GABPB1 sequence variant is conserved across multiple species. In certain embodiments, a plurality of the nucleotide substitutions in the variant are of residues that are conserved across multiple species. In certain embodiments, at least one of the nucleotide substitutions in a peptide variant is of a residue that is conserved among from human, pig, rat, mouse, dog, rabbit, cow, chicken, donkey, sheep, cat, and horse. In certain embodiments, a plurality of the nucleotide substitutions in a variant are of residues that are conserved from human, pig, rat, mouse, dog, rabbit, cow, chicken, donkey, sheep, cat, and horse. [0038] Computational analysis and site-directed mutagenesis experiments in vitro shows that the tetramer form of GABP activates the mutant TERT promoter. This breakthrough approach targets TERT expression in TERTp mutant cells while retaining GABP dimer function. This cancer-specific targeting inhibits the cancer cells while avoiding many normal cell toxicities. [0039] As described herein, nucleic acid-based compositions that “target” one or more regions and/or elements of a GABPB1 gene are designed to hybridize with the transcript of a GABPB1 gene at a specific region, site or location or at a specific element of the transcript. The terms “transcript”, “target transcript” or “target RNA” refer to any RNA transcribed from GABPB1, as the case may be depending on the context, including pre-mRNA and processed mRNA encoding a GABP protein. [0040] Many mechanisms contribute to mRNA stability and translation efficiency. For instance, start codon context (i.e., positions -3 to -1 relative to a start codon of a gene open reading frame), secondary structure of an mRNA, out-of-frame upstream AUGs (uAUGs), and Kozak sequence context have all been implicated in affecting mRNA stability and translation efficiency. Dvir, Shlomi, et al. "Deciphering the rules by which 5′-UTR sequences affect protein expression in yeast." Proceedings of the National Academy of Sciences 110.30 (2013): E2792-E2801; and Li, Jing, et al. "Nucleotides upstream of the Kozak sequence strongly influence gene expression in the yeast S. cerevisiae." Journal of biological engineering 11.1 (2017): 25; each of which is incorporated herein by reference in its entirety. [0041] Specific mechanisms by which the nucleic acid-based compositions described herein can decrease expression of a GABPB1 gene include alleviating the effect of downregulatory cis and trans regulators of GABPB1 translation by enhancing mRNA stability and/or translation efficiency. In some embodiments, nucleic acid-based compounds described herein target binding sites for general and sequence specific translation factors. In some embodiments, nucleic acid-based compositions described herein target sequences implicated in formation of secondary structures that may hamper scanning of the 3′ UTR by translation complexes, thereby alleviating the inhibitory effects of the secondary structures. In some embodiments, nucleic acid-based compositions described herein target sites where RNA binding proteins interact with a GABPB1 mRNA. The nucleic acid-based compositions described herein can, in some embodiments, interrupt the interaction and thereby alleviate downregulatory effects of RNA binding proteins. [0042] The nucleic acid-based compositions described herein may target sequences that form secondary structures involved in RNA decay mechanisms that selectively degrade highly structured RNAs. (See Fischer, Joseph W., et al. "Structure-Mediated RNA Decay by UPF1 and G3BP1." Molecular Cell (2020), incorporated herein by reference in its entirety.) Thus, the nucleic acid-based compositions inhibit structure-mediated RNA decay mechanisms in some embodiments, and thereby increase mRNA stability and translation efficiency. II. Design and Synthesis of Nucleic Acid-Based Compositions: ASOs [0043] Antisense oligonucleotides (ASOs) are typically short (e.g., around 8 – 50 nucleotides), single-stranded oligonucleotide molecules designed to hybridize with an RNA sequence, e.g., mRNA, rRNA. Such targeting may result in either an upregulation or downregulation of the expression of the gene depending on the design of the ASO and the site of action, or localization of the ASO. [0044] ASOs can target and alter messenger RNA to modulate gene expression at the level of translation. (Rinaldi, Carlo, and Matthew JA Wood. Nature Reviews Neurology 14.1 (2018): 9, the contents of which are incorporated by reference herein in their entirety.) ASOs modulate target gene expression through various mechanisms, including RNase H-dependent pathways, ribozyme-based RNA destruction, and steric hindrance of cellular translation machinery. ASOs can also be designed to target and block critical aspects of mRNA processing, such as intron excision and polyadenylation. (Rinaldi and Wood, 2018.) Because of their capacity for broad biodistribution and the fact they can be designed to target virtually any RNA sequence, ASOs have tremendous potential as therapeutics for monogenic diseases where modulation of a single gene product’s expression can ameliorate a disease state or slow or halt disease progression. [0045] Recently, ASOs targeting certain regions, herein referred to as downregulatory sequence elements (DSEs) in 5’ and 3’ untranslated regions (UTRs) have been implemented. For example, Master et al showed that ASOs targeting cis-acting regulatory elements such as secondary structures in 5’ UTRs can be used to increase target gene expression. (Master, Adam, et al. “A Novel Method for Gene-Specific Enhancement of Protein Translation by Targeting 5’ UTRs of Selected Tumor Suppressors.” PloS ONE 11(5): e0155359, the contents of which are incorporated by reference herein in their entirety.) Liang et al showed that ASOs targeting upstream open reading frames (uORFs) improved translation efficiency in targeted genes. (Liang, Xue-hai, et al. "Translation efficiency of mRNAs is increased by antisense oligonucleotides targeting upstream open reading frames." Nature biotechnology 34.8 (2016): 875, the contents of which are incorporated by reference herein in their entirety. See also International Patent Application Publication No. WO2016077837, the contents of which are incorporated by reference herein in their entirety.) Together, these findings open extensive new avenues of investigation for antisense strategies for increasing target gene expression at the level of protein translation. [0046] The ASOs described herein can have an indirect downregulation effect on the RNA transcript(s) transcribed from the template strand of the target gene and/or the polypeptide(s) encoded by the target gene or mRNA. The RNA transcript transcribed from the target gene is referred to as the “target transcript,” “target RNA,” “target mRNA,” or the like, as the case may be. The target transcript can be an mRNA of the target gene. The target transcript can exist in the cytoplasm, the nucleus, or the mitochondrion of a cell. The ASOs of the present disclosure can have a downstream effect on a biological process or activity, including, for example, where targeting a first transcript affects (either by upregulating or downregulating) a second, non-target transcript. [0047] In some embodiments, the ASOs described herein are stable in rat liver microsome. In some embodiments, at least 20% of the ASOs described herein remain in rat liver microsome after 72 hours. In some embodiments, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75% or about 80% of the ASOs described herein remain in rat liver microsome after 72 hours. [0048] In some embodiments, the ASOs described herein, when injected intracranially into mice, do not cause weight loss or clinical signs of weakness/morbidity in mice after 1 week. Targeted Sequence [0049] In some embodiments, the nucleic acid-based compositions described herein, including ASOs described herein, comprise an oligo- or polynucleotide that is at least 80% complementary to a region of the target transcript. This region on the target transcript where the nucleic acid-based compositions hybridize or bind to the target transcript is referred to as the “targeted sequence” or “target site.” [0050] The term “complementary to” means being able to hybridize under physiological conditions, or in some cases being able to hybridize under stringent conditions with respect to hybridization temperature and salt concentration. It is to be understood that thymidine (T) of any given DNA sequence is replaced by uridine (U) in its corresponding RNA transcript and that this difference does not alter the understanding of the term “complementarity.” [0051] The nucleic acid-based compositions described can share at least 80% sequence identity, at least 90% sequence identity, at least 95% sequence identity, at least 98% sequence identity, at least 99% sequence identity, or can be 100% identical with, the reverse complement of the targeted sequence. Thus, the reverse complements of the described nucleic acid-based compositions have a high degree of sequence identity with the targeted sequence. The targeted sequence can have the same length, i.e., the same number of nucleotides, as the nucleic acid based compositions or the targeted sequence can have a similar length i.e., within 1 nucleotide, within 2 nucleotides, within 3 nucleotides, within 4 nucleotides, or within 5 nucleotides compared to the length of the nucleic acid-based compositions. The nucleic acid-based compositions may hybridize with all or a portion of the targeted sequence, or hybridize intermittently with the targeted sequence. In some embodiments, targeted sequence may hybridize with all or a portion of the nucleic acid-based compositions described herein, or the targeted sequence may hybridize intermittently with the nucleic acid- based compositions. [0052] In some embodiments, the targeted sequence comprises at least 8 nucleotides. Thus, the targeted sequence can be 8 nucleotides, 9 nucleotides, 10 nucleotides, 11 nucleotides, 12 nucleotides, 13 nucleotides, 14 nucleotides, 15 nucleotides, 16 nucleotides, 17 nucleotides, 18 nucleotides, 19 nucleotides, 20 nucleotides, 21 nucleotides, 22 nucleotides, 23 nucleotides, 24 nucleotides, 25 nucleotides, 26 nucleotides, 27 nucleotides, 28 nucleotides, 29 nucleotides, or 30 nucleotides in length. In some cases, the targeted sequence is greater than 30 nucleotides in length. In some embodiments, the targeted sequence is between 6 and 18 nucleotides in length. [0053] In some embodiments, the targeted sequence is between 7 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 8 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 9 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 10 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 11 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 12 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 13 and 18 nucleotides in length. In some embodiments, the targeted sequence is between 14 and 18 nucleotides in length. In some embodiments, the targeted sequence is about 14 nucleotides in length. In some embodiments, the targeted sequence is about 15 nucleotides in length. In some embodiments, the targeted sequence is about 16 nucleotides in length. [0054] The targeted sequences described herein can be any sequence derived from a GABPB1 RNA, including a pre-mRNA or an mRNA. References to sequences of GABPB1 are provided in Table 1 and is understood by those of skill in the art that such DNA may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 1. GABPB1 Sequences
Figure imgf000015_0001
[0055] In some embodiments, the targeted sequences described herein are located in a region of GABPB1 such as, but not limited to, an exon, intron or an junction between two exons spanning between 12 nucleotides and 22 nucleotides. For example, the targeted sequence may be 14 nucleotides in length, 15 nucleotides in length, 16 nucleotides in length, 17 nucleotides in length, 18 nucleotides in length, 19 nucleotides in length, 20 nucleotides in length, 21 nucleotides in length, or 22 nucleotides in length. Correspondingly, the GABPB1- hybridizing oligonucleotides described herein can be 14-mers, 15-mers, 16-mers, 17-mers, 18-mers, 19-mers 20-mers, 21-mer, or 22-mers. [0056] In some embodiments, the targeted sequences described herein are located in exon 9 of GABPB1 or the 3’UTR of GABPB1. Non-limiting example of regions or sequences that can be targeted in exon 9 of GABPB1L are described in International Patent Publication No. WO2018217975, the contents of which are herein incorporated by reference in its entirety. In WO2018217975, provided are regions of exon 9 or the 3' untranslated region (UTR) of GABPBIL mRNA which can be targeted by ASOs, and thus can be used to reduce TERT expression and treat cancers harboring TERT promoter mutations. As a non-limiting example, the targeted sequence region of GABPB1L may be, but not limited to, SEQ ID NO. 1-6 and 13-15 as described in WO2018217975, the contents of which are herein incorporated by reference. As a non-limiting example, ASOs which may be used to target exon 9 or the 3' UTR of GABPBIL mRNA are SEQ ID NO: 7-12 of WO2018217975, the contents of which are herein incorporated by reference in its entirety. [0057] As a non-limiting example, targeted sequences that target a sequence within exon 9 of GABPB1 or the intron on the exon 8-exon 9 boundary are provided in Table 2 and Table 3. Nucleic acid-based compositions described herein can be designed to hybridize to a targeted sequence provided in Table 2 and Table 3. Exemplary 15-mer ASOs are provided in Table 2 and SSOs are provided in Table 3. In Table 2 and Table 3, * denotes a phosphorothioate bond in the backbone and + is a LNA. [0058] For all oligonucleotide sequences provided in Table 2 and Table 3, GC content is shown as a percentage (“GCPerc”). By convention, the oligonucleotides are written in the 5’ to 3’ direction and as DNA oligonucleotides. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. [0059] In Table 2 and Table 3 the label “Start” refers to the position on chromosome 15 which corresponds to the start of the region of hybridization of the hybridizing oligonucleotide sequences and also to the first nucleotide of the targeted region shown in the table as the “GABP Targeted Sequence,” and “End” refers to the position on chromosome 15 which corresponds to the end of the region of hybridization. Hybridization of the target sequences described herein may be at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 99%, or 100%. For example, in Table 2, the GABPB1-hybridizing oligonucleotide of SEQ ID NO: 19 will hybridize to the region on chromosome 15 from Start to End of nucleotide 50570781 to 50570796. Likewise, SEQ ID NO: 24166 is the actual sequence from this region on chromosome 15 to which the oligonucleotide hybridizes. Also provided in Tables 2 and 3 are the sequence identifier for the unmodified ASO sequence (“Unmod. SEQ ID”). Table 2. GABPB115-mer Targeted Sequences and ASO Sequences
Figure imgf000016_0001
Figure imgf000017_0001
Figure imgf000018_0001
Figure imgf000019_0001
[0060] In some embodiments, the GABPB1-hybridizing sequence comprises SEQ ID NO: 38 or at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 38. In some embodiments, the GABPB1-hybridizing sequence comprises SEQ ID NO: 39 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 39. In some embodiments, the GABPB1 -hybridizing sequence comprises SEQ ID NO: 54 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 54. In some embodiments, the GABPBl- hybridizing sequence comprises SEQ ID NO: 57 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 57. In some embodiments, the GABPBl-hybridizing sequence comprises SEQ ID NO: 433 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 433. In some embodiments, the GABPB1 -hybrbdizing sequence comprises SEQ ID NO: 450 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 450.
Table 3. GABPB1 15-mer Targeted Sequences and SSO Sequences is
Figure imgf000020_0001
Figure imgf000021_0001
Figure imgf000022_0001
Figure imgf000023_0001
GABPB1 Hybridizing Oligonucleotides [0061] As discussed above, the nucleic acid-based compositions described have a high degree of sequence identity with the targeted sequence of the RNA transcript. Instead of "complementary to the targeted sequence," the nucleic acid-based compositions can also be defined as having "identity" to a region on the coding strand of the target gene. Therefore, the genomic sequence of the target gene may be used to design nucleic acid-based compositions. These compositions, referred to herein as “GABPB1-hybridizing oligonucleotides” will thus hybridize with the targeted sequence of the RNA transcript The targeted sequence may be the GABPB1L isoform, the GABPB1S isoform or may target both isoforms for a reduction in total GABPB 1.
[0062] Off-target hits of the nucleic acid-based compositions designed as described herein can be determined. The sequences targeted by the nucleic acid-based compositions are compared against the target transcriptome represented by the human RefSeq database. A number N of mismatches can be permitted in the identification of off-target hits. Off-target mismatch hit number refers to the number of nucleotide mismatches permitted in an off- target hit when comparing the targeted sequence to off-target hit sequences in the transcriptome. Off-target hits in the transcriptome are counted and 0 mismatch hit number and 1 mismatch hit number off-target hits are then determined. For example, the term “M0” refers to the number of unique genes with transcripts other than the target transcripts of the nucleic acid-based composition which the nucleic acid-based composition may hybridize with or bind to with 0 mismatched base pairs. In other words, “M0” counts the number of unique genes among known RefSeq transcripts in the target genome, other than the target transcript, that comprise a region completely identical with the complement of the nucleic acid-based composition sequence. In some embodiments, only nucleic acid-based compositions having no off-target hits defined as no M0 hits and no Ml hits are selected for further use or testing.
[0063] Non-limiting examples of nucleic acid-based compositions described herein include GABPB 1 -hybridizing oligonucleotides comprising about 14 nucleotides, about 15 nucleotides, or about 16 nucleotides. The sequence of GABPB1 -hybridizing oligonucleotides can have at least 60%, 70%, 80% or 90% identity to a reverse complement of a targeted sequence selected from or derived from the exon of GABPB1 mRNA or a sequence provided in Table 2 or Table 3. The sequence of GABPB 1 -hybridizing oligonucleotides can have at least 60%, 70%, 80% or 90% identity to a reverse complement of a targeted sequence selected from or derived from the exon of GABPB 1 mRNA, intron of GABPB 1 mRNA or a sequence provided in Table 2, Table 3, Table 4, Table 5, Table 6, Table 7 or Table 8. In some embodiments, the nucleic acid-based compositions comprise a sequence selected from the sequences of Table 2, Table 3, Table 4, Table 5, Table 6, Table 7 or Table 8. In some embodiments, the GABPB 1 -hybridizing oligonucleotides have a 3’ tail.
[0064] Non-limiting examples of nucleic acid-based compositions described herein include GABPB 1 -hybridizing oligonucleotides comprising about 14 nucleotides, about 15 nucleotides, about 16 nucleotides, about 17 nucleotides, about 18 nucleotides, about 19 nucleotides, or about 20 nucleotides. The sequence of GABPB 1 -hybridizing oligonucleotides can have at least 60%, 70%, 80% or 90% identity to a sequence selected from or derived from the exon of GABPB1 mRNA or a sequence provided in Table 2, Table 3, Table 4, Table 5, Table 6, Table 7, or Table 8. In some embodiments, the nucleic acid-based compositions comprise a sequence selected from the sequences of Table 2, Table 3, Table 4, Table 5, Table 6, Table 7, or Table 8. In some embodiments, the GABPB1-hybridizing oligonucleotides have a 3’ tail. [0065] GABPB1-hybridizing oligonucleotides can be unmodified, or can be modified according to the modifications described herein. The GABPB1-hybridizing oligonucleotides can be fully modified. In some embodiments, GABPB1-hybridizing oligonucleotides are partially modified. [0066] The nucleic acid-based compositions of the present disclosure can be produced by any suitable method, for example synthetically or by expression in cells using standard molecular biology techniques which are well-known to a person of ordinary skill in the art. For example, ASOs can be chemically synthesized or recombinantly produced using methods known in the art. Antisense Oligonucleotides [0067] Some embodiments of the nucleic acid-based compositions described herein are antisense oligonucleotides (ASOs). In some embodiments, an ASO of the present disclosure is a single-stranded oligonucleotide. In some embodiments, the ASO is a single-stranded oligodeoxynucleotide. The length of an ASO determines its biodistribution upon administration. Thus, in some instances, the length of the ASO is optimized for biodistribution to the CNS, including but not limited to the brain. As a non-limiting example, the ASOs may be optimized to distribute to areas of the brain that have been effected by tumor growth as a result of glioblastoma or other brain tumors. . As such, the ASO can be between about 8 and about 25 nucleotides in length. In some embodiments, an ASO of the present disclosure is at least 8, at least 9, at least 10, at least 11, 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 nucleotides in length. In some embodiments, an ASO is 8 nucleotides in length, 9 nucleotides in length, 10 nucleotides in length, 11 nucleotides in length, 12 nucleotides in length, 13 nucleotides in length, 14 nucleotides in length, 15 nucleotides in length, 16 nucleotides in length, 17 nucleotides in length, 18 nucleotides in length, 19 nucleotides in length, 20 nucleotides in length, 21 nucleotides in length, 22 nucleotides in length, 23 nucleotides in length, 24 nucleotides in length, 25 nucleotides in length or more than 25 nucleotides in length In some embodiments an ASO is up to about 30 nucleotides in length. Preferably, the ASO of the present disclosure has a length of about 14 nucleotides, or has a length of exactly 14 nucleotides, so as to promote biodistribution to cells of the kidney upon administration to a subject, to promote biodistribution to cells of the liver upon administration to a subject, or to promote biodistribution to cells of the kidney and cells of the liver upon administration to a subject. In some embodiments, the ASO has at least 80%, 90%, 95%, 98%, 99% or 100% complementarity to the targeted sequence.
[0068] The ASOs described herein have a high degree of complementarity to the targeted sequence of the RNA transcript. In some embodiments, the ASO has no more than 5, no more than 4, no more than 3, no more than 2, no more than 1, or no mismatches with reference to the targeted sequence of the RNA transcript.
[0069] A “strand” in the context of the present disclosure means a contiguous sequence of nucleotides, including non-naturally occurring or modified nucleotides. Two or more strands may be, or each form a part of, separate molecules, or they may be connected covalently, e.g., by a linker such as a polyethylene glycol linker.
[0070] Some embodiments of ASOs as described herein involve formation of a duplex. An ASO duplex refers to a single ASO molecule that is at least partly self-complementary/ and capable of forming a hairpin structure, including a duplex region. In such case, the term “strand” refers to one of the regions of the ASO that is complementary to another internal region of the ASO. In such embodiments, typically, one strand will target the targeted sequence (the “guide strand”) and the other strand will act merely as a “passenger strand.” The guide strand of the ASO will have no more than 5, or no more than 4 or 3, or no more than 2, or no more than 1, or no mismatches with the targeted sequence of the RNA transcript.
[0071] In some embodiments, the passenger strand of a duplex ASO comprises at least one nucleotide that is not complementary/ to the corresponding nucleotide on the guide strand, called a mismatch with the guide strand. The at least one mismatch with the guide strand can be at 3’ end of the passenger strand . In some embodiments, the 3’ end of the passenger strand comprises 1-5 mismatches with the guide strand. In some embodiments, the 3’ end of the passenger strand comprises 2-3 mismatches with the guide strand. In some embodiments, the 3’ end of the passenger strand comprises 6-10 mismatches with the guide strand.
[0072] A duplex ASO can have siRNA-like complementarity to the targeted sequence on the template strand; that is, 100% complementarity between nucleotides 2-6 from the 5’ end of the guide strand and a region of the targeted sequence. Other nucleotides of the ASO can, in addition, have at least 80%, 90%, 95%, 98%, 99% or 100% complementarity to a region of the targeted sequence. For example, nucleotides 7 (counted from the 5' end) until the 3' end of the ASO can have least 80%, 90%, 95%, 98%, 99% or 100% complementarity to a region of the targeted sequence. [0073] The terms “small interfering RNA” or “siRNA” mean a double-stranded RNA typically 20-25 nucleotides long involved in the RNA interference (RNAi) pathway and interfering with or inhibiting the expression of a specific gene. The gene is the target gene of the siRNA. A siRNA is usually about 21 nucleotides long, with 3' overhangs (e.g., 2 nucleotides) at each end of the two strands. A siRNA inhibits target gene expression by binding to and promoting the cleavage of one or more RNA transcripts of the target gene at specific sequences. Typically, in RNAi the RNA transcripts are mRNA, so cleavage of mRNA results in the down-regulation of gene expression. [0074] In some embodiments, the ASO comprises a number of unpaired nucleotides at the 3' end forming 3' overhangs or tails. The number of unpaired nucleotides forming the 3' overhang can range from 0 to 5 nucleotides, or 1 to 3 nucleotides, or 2 nucleotides. [0075] Thus, in some embodiments, the ASOs of the present disclosure consist of (i) a sequence having at least 80% complementarity to a targeted sequence of the target transcript; and (ii) a 3' tail of 1-5 nucleotides, which may comprise uracil residues, such as UU, UUU, or mUmU (m stands for 2’-OMe modification). In some embodiments, the ASOs of the present disclosure are duplex ASOs having a double-stranded duplex region, wherein a guide strand comprises (i) a first sequence having at least 80% complementarity to a targeted sequence; (ii) a 3' overhang of 1-5 nucleotides and a passenger strand that forms a duplex with the guide sequence. Any 3’ tail shall not be regarded as contributing to mismatches for purposes of determining complementarity between the ASO and the targeted sequence. [0076] The ASOs of the present disclosure can contain a flanking sequence. The flanking sequence can be at the 3’ end or 5’ end of the ASO. In some embodiments, the flanking sequence is the sequence of a miRNA, conferring on the ASO a miRNA configuration which may be processed with Drosha and Dicer. In a non-limiting example, an ASO of the present disclosure has two strands and is cloned into a microRNA precursor, e.g., miR-30 backbone flanking sequence. [0077] The ASOs of the present disclosure can comprise a restriction enzyme substrate or recognition sequence. The restriction enzyme recognition sequence can be at the 3’ end or 5’ end of the ASO. Non-limiting examples of restriction enzymes include NotI and AscI. Sterochemically Controlled ASOs Chirally Controlled ASOs [0078] The stereochemical designation of the chiral linkage in the ASOs of the present disclosure may be controlled using methods know in the art. As a non-limiting example, the ASOs of the present disclosure may be chirally controlled oligonucleotides such as those described in International Patent Publication No. WO2014012081, the contents of which are herein incorporated by reference. [0079] In some embodiments, the present disclosure provides chirally controlled ASOs, and chirally controlled ASO compositions which are of high crude purity. In some embodiments, the present disclosure provides chirally controlled ASOs, and chirally controlled ASO compositions which are of high diastereomeric purity. [0080] In some embodiments, the present disclosure provides chirally controlled compositions comprising a plurality of ASOs of at least one type, wherein each type is defined by: 1) base sequence; 2) pattern of backbone linkages; 3) pattern of backbone chiral centers; and 4) pattern of backbone P-modifications. In some embodiments, the present disclosure provides chirally controlled compositions comprising a plurality of ASOs of the same type, wherein each type is defined by: 1) base sequence; 2) pattern of backbone linkages; 3) pattern of backbone chiral centers; and 4) pattern of backbone P-modifications. In some embodiments, the present disclosure provides chirally controlled compositions comprising a plurality of oligonucleotides of two or more types, wherein each type is defined by: 1) base sequence; 2) pattern of backbone linkages; 3) pattern of backbone chiral centers; and 4) pattern of backbone P-modifications. [0081] In some embodiments, the present disclosure provides ASOs comprising one or more diastereomerically pure internucleotidic linkages with respect to the chiral linkage phosphorus. In some embodiments, the present disclosure provides ASOs comprising one or more diastereomerically pure internucleotidic linkages having the structure of formula I as described in WO2014012081, the contents of which are herein incorporated by reference in their entirety. In some embodiments, the present disclosure provides ASOs comprising one or more diastereomerically pure internucleotidic linkages with respect to the chiral linkage phosphorus, and one or more phosphate diester linkages. In some embodiments, the present disclosure provides oligonucleotides comprising one or more diastereomerically pure internucleotidic linkages having the structure of formula I of WO2014012081, and one or more phosphate diester linkages. In some embodiments, the present disclosure provides ASOs comprising one or more diastereomerically pure internucleotidic linkages having the structure of formula I-c of WO2014012081, and one or more phosphate diester linkages. Exemplary internucleotidic linkages, including those having structures of formula I of WO2014012081, are further described below. In some embodiments, such oligonucleotides comprise a sequence further described in the application, including but not limited to those described in Tables 2 and 4, and Appendices A, B and C of WO2014012081. [0082] In some embodiments, a provided ASO comprises a combination of stereopure and stereorandom internucleotidic linkages with respect to chirality at the linkage phosphorus. For instance, in some embodiments it is desirable to have a block of one or more stereodefmed internucleotidic linkages within an ASO that is otherwise stereorandom with respect to chirality at the linkage phosphorus. In some embodiments, it is desirable to have a block of one or more internucleotidic linkages that are stereorandom within an ASO that is otherwise stereodefined with respect to chirality at the linkage phosphorus. [0083] In some embodiments, at least one nucleotide unit of a provided ASO is installed using stereoselective oligonucleotide synthesis, as described in this application, to form a pre- designed diastereomerically pure internucleotidic linkage with respect to the chiral linkage phosphorus. In some embodiments, at least two nucleotide units of a provided ASO are installed using stereoselective oligonucleotide synthesis, as described in this application, to form at least two pre-designed diastereomerically pure internucleotidic linkages with respect to the chiral linkage phosphorus. In some embodiments, at least three nucleotide units of a provided ASO are installed using stereoselective oligonucleotide synthesis, as described in WO2014012081, to form at least three pre-designed diastereomerically pure internucleotidic linkages with respect to the chiral linkage phosphorus. In some embodiments, the at least one, two, or three pre-designed diastereomerically pure internucleotidic linkages are adjacent to one another. In some embodiments, the at least one, two, or three pre-designed diastereomerically pure internucleotidic linkages are not adjacent to one another. [0084] In some embodiments, at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% of nucleotide units of a provided ASO are installed using stereoselective oligonucleotide synthesis, as described in this application, to form a pre-designed diastereomerically pure internucleotidic linkage with respect to the chiral linkage phosphorus. As described herein, in some embodiments the at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% of nucleotide units occur in one or more blocks to provide a blockmer. In some embodiments, the at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% of nucleotide units occur in an alternating pattern to provide an altmer. One of skill in the relevant arts will recognize that any desirable pattern can be achieved using methods of the present disclosure and are contemplated herein. [0085] In some embodiments, the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different stereochemistry and/or different P-modifications relative to one another. In certain embodiments, the present disclosure provides a chirally controlled ASO, wherein at least two individual internucleotidic linkages within the ASO have different P-modifications relative to one another. In certain embodiments, the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different P-modifications relative to one another, and wherein the chirally controlled ASO comprises at least one phosphate diester internucleotidic linkage. In certain embodiments, the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different P-modifications relative to one another, and wherein the chirally controlled ASO comprises at least one phosphate diester internucleotidic linkage and at least one phosphorothioate diester internucleotidic linkage. In certain embodiments, the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different P- modifications relative to one another, and wherein the chirally controlled ASO comprises at least one phosphorothioate triester internucleotidic linkage. In certain embodiments, the present disclosure provides a chirally controlled ASO, wherein at least two of the individual internucleotidic linkages within the ASO have different P-modifications relative to one another, and wherein the chirally controlled ASO comprises at least one phosphate diester internucleotidic linkage and at least one phosphorothioate triester internucleotidic linkage. [0086] Stereoisomers of a particular oligonucleotide can show different stability and/or activity (e.g., functional and/or toxicity properties) from each other. As shown in WO2017015555, the contents of which are herein incorporated by reference in their entirety, stability and/or activity improvements may be achieved through inclusion and/or location of particular chiral structures within an oligonucleotide can be comparable to, or even better than those achieved through use of particular backbone linkages, residue modifications, etc. (e.g., through use of certain types of modified phosphates (e.g., phosphorothioate, substituted phosphorothioate, etc.), sugar modifications (e.g., 2’- modifications, etc.), and/or base modifications (e.g., methylation, etc.)). [0087] In some embodiments, properties (e.g., stability and/or activities) of an ASO can be adjusted by optimizing its pattern of backbone chiral centers, optionally in combination with adjustment/optimization of one or more other features (e.g., linkage pattern, nucleoside modification pattern, etc.) of the ASO as described in WO2017015555, the contents of which are herein incorporated by reference in their entirety. [0088] In some embodiments, the present disclosure provides compositions of ASOs, wherein the ASOs have a common pattern of backbone chiral centers which may enhance the stability and/or biological activity of the ASOs. In some embodiments, a pattern of backbone chiral centers provides increased stability. In some embodiments, a pattern of backbone chiral centers provides increased activity. In some embodiments, a pattern of backbone chiral centers provides increased stability and activity. In some embodiments, a pattern of backbone chiral centers, such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, effectively prevents cleavage at secondary sites. In some embodiments, a pattern of backbone chiral centers , such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, creates new cleavage sites. In some embodiments, a pattern of backbone chiral centers, such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, minimizes the number of cleavage sites. In some embodiments, a pattern of backbone chiral centers, such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, minimizes the number of cleavage sites so that a target nucleic acid polymer is cleaved at only one site within the sequence of the target nucleic acid polymer that is complementary to the ASO. In some embodiments, a pattern of backbone chiral centers, such as those described in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, enhances cleavage efficiency at a cleavage site. In some embodiments, a pattern of backbone chiral centers of the ASO improves cleavage of a target nucleic acid polymer. In some embodiments, a pattern of backbone chiral centers increases selectivity. In some embodiments, a pattern of backbone chiral centers minimizes off-target effect. In some embodiments, a pattern of backbone chiral centers increase selectivity, e.g., cleavage selectivity between two target sequences differing only by a single nucleotide polymorphism (SNP). Exemplary backbone chiral centers are provided in WO2017015555 the contents of which are herein incorporated by referenced in their entirety, [0089] In some embodiments, the present disclosure provides compositions and methods for altering splicing of transcripts. Splicing of a transcript, such as pre-mRNA, is an essential step for the transcript to perform its biological functions in many higher eukaryotes. Defects d/ i ffi i i h li i ff bi l i l f i d/ h pathological consequences. For example, many human genetic diseases are caused by mutations that cause splicing defects, and many diseases are associated with splicing defects that are not attributed to overt mutations. In some embodiments, the present disclosure recognizes that targeting splicing, especially through compositions comprising oligonucleotides having the chemical modifications and/or stereochemistry patterns described in this disclosure, can effectively correct disease-associated aberrant splicing, and/or introduce and/or enhance beneficial splicing that lead to desired products, e.g., mRNA, proteins, etc. which can repair, restore, or add new desired biological functions. For example, in some embodiments, by skipping one or more exons of a pre-mRNA to produce an mRNA with frameshift and/or premature termination codon, provided compositions and methods effectively knockdown a gene; in some embodiments, such a gene is a mutant gene. [0090] In some embodiments, the ASOs of the present disclosure may include chemical modifications, stereochemistry and combinations thereof can be used to modulate splicing of transcripts. Provided in International Patent Publication WO2017062862 and WO2018067973, the contents of each of which are herein incorporated by reference in their entirety, are chemical modifications and patterns thereof useful for improving transcript splicing by oligonucleotides. Among other things, WO2017062862 and WO2018067973 describes how stereochemistry can be used to modulate transcript splicing by oligonucleotide compositions which can be used with the ASO compositions described herein. In some embodiments, combinations of chemical modifications and stereochemistry may be used to improve properties of oligonucleotides, e.g., their capabilities to alter splicing of transcripts. In some embodiments, chirally controlled oligonucleotide compositions that, when compared to a reference condition (e.g., absence of the composition, presence of a reference composition (e.g., a stereorandom composition of oligonucleotides having the same base sequence, the same chemical modifications, etc., a chirally controlled oligonucleotide composition of another stereoisomer, etc.), and combinations thereof), provide altered splicing that can deliver one or more desired biological effects, for example, increase production of desired proteins, knockdown of a gene by producing mRNA with frameshift mutations and/or premature termination codons, knockdown of a gene expressing a mRNA with a frameshift mutation and/or premature termination codon, etc. [0091] In some embodiments, the stereochemistry of an ASO described herein may be modified to improve the properties of the ASO in order to increase the effectiveness of the ASO to the target gene. As a non-limiting example, the stereochemistry of the ASO may be modified as described in International Patent Publication Nos. WO2018223081, WO2018223073, WO2018223056, WO2019200185, the contents of each of which are herein incorporated by reference in their entirety, where the stereochemistry of the backbone chiral centers were modified to increase the properties of the oligonucleotides to specific target genes. Pohosphorodiamidate Morpholino ASOs [0092] ASOs of the present disclosure may be prepared using the stereospecific synthesis of diastereomerically pure or substantially diastereomerically pure phosphorodiamidate morpholino oligomers (PMOs) of US Patent Publication No. US20200115405, the contents of which are herein incorporated by reference, which include at least one chiral phosphorous linkage. Stereochemically pure or substantially stereochemically pure monomers may be prepared by the separation of a diasteromeric mixture of monomers as described in US Patent Publication No. US20200115405, the contents of which are herein incorporated by reference, in paragraphs 48-56, and 62-102. Additionally, the activated monomers may be used to accomplish stereospecific coupling for the preparation of stereochemically pure dinucleotides, stereochemically pure trinucleotides, and larger stereochemically pure oligomers as described in paragraphs 58-61, and 103-161 of US Patent Publication No. US20200115405, the contents of which are herein incorporated by reference. Exemplary ASO Sequences [0093] Exemplary 16-mer and 20-mer ASOs are provided in Table 4. In Table 4, * denotes a phosphorothioate bond in the backbone, + is a LNA, “/52MOErN/” is a 5' 2-MOE base, “/32MOErN/” is a 3' 2-MOE base, “/i2MOErN/” is an internal 2-MOE base, and “/iMe- dC/” is a 5-methyl dC base. Also provided in Table 4 is the name and sequence identifier for the unmodified ASO sequence (“Unmod. ASO Name” and “Unmod. SEQ ID”) and the sequence identifier for the target sequence (“Target SEQ ID”). By convention, the oligonucleotides are written in the 5’ to 3’ direction and as DNA oligonucleotides. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 4. GABPB116-mer and 20-mer ASO Sequences
Figure imgf000033_0001
Figure imgf000034_0001
Figure imgf000035_0001
Figure imgf000036_0001
[0094] Exemplary ASOs are provided in Table 5 and Table 6 which target exon 9, the intron between exon 8 and exon 9, the exon 8-exon 9 junction or total GABPB1 (shown in the table in “Target Area”). Also provided are the sequence identifiers for the target sequence for the ASOs (“Target SEQ ID”). In Table 5, “Int.E8-E9” refers to the intron between exon 8 and 9, “E9” is exon 9 and “Jun.E8-E9” is the exon 8-exon 9 junction. In Table 6, “Total” refers to targeting total GABPB1 (both GABPB1S and GABPB1L) expression. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 5. ASOs Targeting GABPB1
Figure imgf000036_0002
Figure imgf000037_0001
Figure imgf000038_0001
Figure imgf000039_0001
Figure imgf000040_0001
Figure imgf000041_0001
Figure imgf000042_0001
Figure imgf000043_0001
Figure imgf000044_0001
Figure imgf000045_0001
Figure imgf000046_0001
Figure imgf000047_0001
Figure imgf000048_0001
Figure imgf000049_0001
Figure imgf000050_0001
Figure imgf000051_0001
Figure imgf000052_0001
Figure imgf000053_0001
Figure imgf000054_0001
Figure imgf000055_0001
Figure imgf000056_0001
Figure imgf000057_0001
Figure imgf000058_0001
Figure imgf000059_0001
Figure imgf000060_0001
Figure imgf000061_0001
Figure imgf000062_0001
Figure imgf000063_0001
Figure imgf000064_0001
Figure imgf000065_0001
Figure imgf000066_0001
Figure imgf000067_0001
Figure imgf000068_0001
Figure imgf000069_0001
Figure imgf000070_0001
Figure imgf000071_0001
Figure imgf000072_0001
Figure imgf000073_0001
Figure imgf000074_0001
Figure imgf000075_0001
Figure imgf000076_0001
Figure imgf000077_0001
Figure imgf000078_0001
Figure imgf000079_0001
Figure imgf000080_0001
Figure imgf000081_0001
Figure imgf000082_0001
Figure imgf000083_0001
Figure imgf000084_0001
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Figure imgf000086_0001
Figure imgf000087_0001
Figure imgf000088_0001
Figure imgf000089_0001
Figure imgf000090_0001
Figure imgf000091_0001
Figure imgf000092_0001
Figure imgf000093_0001
Figure imgf000094_0001
Figure imgf000095_0001
Figure imgf000096_0001
Figure imgf000097_0001
Figure imgf000098_0001
Figure imgf000099_0001
Figure imgf000100_0001
Figure imgf000101_0001
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Figure imgf000105_0001
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Figure imgf000107_0001
Figure imgf000108_0001
Figure imgf000109_0001
Figure imgf000110_0001
Figure imgf000111_0001
Figure imgf000112_0001
Figure imgf000113_0001
Figure imgf000114_0001
Figure imgf000115_0001
Figure imgf000116_0001
Figure imgf000117_0001
Figure imgf000118_0001
Figure imgf000119_0001
Figure imgf000120_0001
Figure imgf000121_0001
Figure imgf000122_0001
Figure imgf000123_0001
Figure imgf000124_0001
Figure imgf000125_0001
Figure imgf000126_0001
Figure imgf000127_0001
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Figure imgf000131_0001
Figure imgf000132_0001
Figure imgf000133_0001
Figure imgf000134_0001
Figure imgf000135_0001
Figure imgf000136_0001
Table 6. ASOs Targeting Total GABPB1
Figure imgf000136_0002
Figure imgf000137_0001
Figure imgf000138_0001
Figure imgf000139_0001
Figure imgf000140_0001
Figure imgf000141_0001
Figure imgf000142_0001
Figure imgf000143_0001
Figure imgf000144_0001
Figure imgf000145_0001
Figure imgf000146_0001
Figure imgf000147_0001
Figure imgf000148_0001
[0095] Exemplary 20-mer ASOs are provided in Table 7. Also provided are the sequence identifiers for the target sequence for the ASOs (“Target SEQ ID”). In Table 7, “dN” is a DNA base, “(5MdC)” is a 5-Methyl-dC base, “Nm” is a MOE base, and “s” is a phosphorothioate bond. By convention, the oligonucleotides are written in the 5’ to 3’ direction and as DNA oligonucleotides. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 7. GABPB120-mer ASO Sequences
Figure imgf000148_0002
Figure imgf000149_0001
Figure imgf000150_0001
Figure imgf000151_0001
Figure imgf000152_0001
Figure imgf000153_0001
[0096] Exemplary 16-mer, 18-mer and 20-mer ASOs are provided in Table 8 (sequence identifier provided under ASO SEQ ID (with mods)) along with the identifier for the sequence without modifications (ASO SEQ ID (no mods)) and the target sequence (Target SEQ ID) . In Table 8, “dN” is a DNA base, “(5MdC)” is a 5-Methyl-dC base, “Nm” is a MOE base, N+ is a LNA and “s” is a phosphorothioate bond. By convention, the oligonucleotides are written in the 5’ to 3’ direction and as DNA oligonucleotides. It is understood by those of skill in the art that such oligonucleotides may be converted to RNA counterparts, where “T” would be replaced with “U”. Table 8. 16-mer, 18-mer and 20-mer ASO Sequences
Figure imgf000154_0001
Figure imgf000155_0001
Figure imgf000156_0001
[0097] In some embodiments, the ASO sequence comprises SEQ ID NO: 12335 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12335. In some embodiments, the ASO sequence comprises SEQ ID NO: 12338 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12338. In some embodiments, the ASO sequence comprises SEQ ID NO: 12363 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12363. In some embodiments, the ASO sequence comprises SEQ ID NO: 12372 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12372. In some embodiments, the ASO sequence comprises SEQ ID NO: 12376 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12376. In some embodiments, the ASO sequence comprises SEQ ID NO: 12379 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12379. In some embodiments, the ASO sequence comprises SEQ ID NO: 12382 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12382. In some embodiments, the ASO sequence comprises SEQ ID NO: 12383 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12383. In some embodiments, the ASO sequence comprises SEQ ID NO: 12384 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12384. In some embodiments, the ASO sequence comprises SEQ ID NO: 12389 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12389. In some embodiments, the ASO sequence comprises SEQ ID NO: 12393 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12393. In some embodiments, the ASO sequence comprises SEQ ID NO: 12394 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12394. In some embodiments, the ASO sequence comprises SEQ ID NO: 12395 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12395. In some embodiments, the ASO sequence comprises SEQ ID NO: 12413 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12413. In some embodiments, the ASO sequence comprises SEQ ID NO: 12421 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12421. In some embodiments, the ASO sequence comprises SEQ ID NO: 12422 or is at least 75%, 80%, 85%, 90%, 95%, or 99% identical to SEQ ID NO: 12422. [0098] In some embodiments, a selectivity index (SI), which uses the cell viability and knockdown percentages, may be used to compare a selection of ASOs and may be used as a component in determining which of the ASOs have the higher chance of success for additional studies and ultimately to be use and the compositions and methods described herein. The formula for the SI is: SI = Viability % / Knockdown %. Exemplary ASO duplex Sequences [0099] Exemplary ASO duplexes of the present disclosure are provided in Table 9. These ASO duplexes may be used as siRNA duplexes or the individual sequences or fragments thereof may be used as siRNA sequences. The target sequence and target sequence identifier (“TSID”), GC Percentage (“GC %”), the sense sequence and sense sequence identifier (“SSID”), and antisense sequence and antisense sequence identifier (“ASID”) are all shown in Table 9. Table 9. ASO Duplex Sequences
Figure imgf000158_0001
Figure imgf000159_0001
Figure imgf000160_0001
Bifunction Oligonucleotides [0100] Bifunctional or dual-functional nucleic acid-based compositions, including ASOs, can be designed to up-regulate the expression of a first gene and down-regulate the expression of at least one second gene. One portion of the dual-functional nucleic acid-based compositions activate the expression of the first gene and another portion inhibits the expression of the second gene. Chemical Modifications [0101] The terms “modification” or, as appropriate, “modified” refer to structural and/or chemical modifications with respect to A, G, T/U or C nucleotides. Nucleotides in the nucleic acid-based compositions of the present disclosure can comprise non-standard nucleotides, such as non-naturally occurring nucleotides or chemically synthesized nucleotides or deoxynucleotides. [0102] As described herein “nucleoside” is defined as a compound containing a sugar molecule (e.g., a pentose or ribose) or a derivative thereof in combination with an organic base (e.g., a purine or pyrimidine) or a derivative thereof (also referred to herein as “nucleobase”). As described herein, “nucleotide” is defined as a nucleoside including a phosphate group or other backbone linker (internucleoside linkage). [0103] The nucleic acid-based compositions of the present disclosure can include any useful modification, such as to the sugar, the nucleobase, or the internucleoside linkage (e.g. to a linking phosphate / to a phosphodiester linkage / to the phosphodiester backbone). One or more atoms of a pyrimidine nucleobase may be replaced or substituted with optionally substituted amino, optionally substituted thiol, optionally substituted alkyl (e.g., methyl or ethyl), or halo (e.g., chloro or fluoro). In certain embodiments, modifications (e.g., one or more modifications) are present in each of the sugar and the internucleoside linkage. Modifications according to the present disclosure may be modifications of ribonucleic acids (RNAs) to deoxyribonucleic acids (DNAs), threose nucleic acids (TNAs), glycol nucleic acids (GNAs), peptide nucleic acids (PNAs), locked nucleic acids (LNAs) or hybrids thereof. In some embodiments, the modifications include 2’-OMe-modified or 2’-O-(2- Methoxyethyl)-modified nucleotides (2’OMe and 2’MOE modifications, respectively). [0104] In some embodiments, the nucleic acid-based compositions of the present di l i l difi i d ib d h i [0105] The nucleic acid-based compositions of the present disclosure can include a combination of modifications to the sugar, the nucleobase, and/or the internucleoside linkage. [0106] The nucleic acid-based compositions of the present disclosure may be modified with any modifications of an oligonucleotide or polynucleotide disclosed in PCT Publication WO2013/151670, the contents of which are incorporated herein by reference in their entirety, including modifications to one or more sugar, to one or more base, or to one or more internucleoside linkage of a an oligonucleotide. Modified Nucleobases [0107] The modified nucleotide base pairing may encompass not only the standard adenosine-thymine, adenosine-uracil, or guanosine-cytosine base pairs, but also base pairs formed between nucleotides and/or modified nucleotides comprising non-standard or modified bases, wherein the arrangement of hydrogen bond donors and hydrogen bond acceptors permits hydrogen bonding between a non-standard base and a standard base or between two complementary non-standard base structures. One example of such non- standard base pairing is the base pairing between the modified nucleotide inosine and adenine, cytosine or uracil. [0108] The modified nucleosides and nucleotides can include a modified nucleobase. Examples of nucleobases found in RNA include, but are not limited to, adenine, guanine, cytosine, and uracil. Examples of nucleobase found in DNA include, but are not limited to, adenine, guanine, cytosine, and thymine. [0109] In some embodiments, the modified nucleobase is a modified uracil. Exemplary nucleobases and nucleosides having a modified uracil include pseudouridine (ψ), pyridin-4- one ribonucleoside, 5-aza-uridine, 6-aza-uridine, 2-thio-5-aza-uridine, 2-thio-uridine (s2U), 4- thio-uridine (s4U), 4-thio-pseudouridine, 2-thio-pseudouridine, 5-hydroxy-uridine (ho5U), 5- aminoallyl-uridine, 5-halo-uridine (e.g., 5-iodo-uridine or 5-bromo-uridine), 3-methyl-uridine (m3U), 5-methoxy-uridine (mo5U), uridine 5-oxyacetic acid (cmo5U), uridine 5-oxyacetic acid methyl ester (mcmo5U), 5-carboxymethyl-uridine (cm5U), 1-carboxymethyl- pseudouridine, 5-carboxyhydroxymethyl-uridine (chm5U), 5-carboxyhydroxymethyl-uridine methyl ester (mchm5U), 5-methoxycarbonylmethyl-uridine (mcm5U), 5- methoxycarbonylmethyl-2-thio-uridine (mcm5s2U), 5-aminomethyl-2-thio-uridine (nm5s2U), 5-methylaminomethyl-uridine (mnm5U), 5-methylaminomethyl-2-thio-uridine (mnm5s2U), 5- methylaminomethyl-2-seleno-uridine (mnm5se2U), 5-carbamoylmethyl-uridine (ncm5U), 5- carboxymethylaminomethyl-uridine (cmnm5U), 5-carboxymethylaminomethyl-2-thio-uridine (cmnm5s2U), 5-propynyl-uridine, 1-propynyl-pseudouridine, 5-taurinomethyl-uridine (τm5U), 1-taurinomethyl-pseudouridine, 5-taurinomethyl-2-thio-uridine(τm5s2U), 1-taurinomethyl-4- thio-pseudouridine, 5-methyl-uridine (m5U, i.e., having the nucleobase deoxythymine), 1- methylpseudouridine (m1ψ), 5-methyl-2-thio-uridine (m5s2U), 1-methyl-4-thio-pseudouridine (m1s4ψ), 4-thio-1-methyl-pseudouridine, 3-methyl-pseudouridine (m3ψ), 2-thio-1-methyl- pseudouridine, 1-methyl-1-deaza-pseudouridine, 2-thio-1-methyl-1-deaza-pseudouridine, dihydrouridine (D), dihydropseudouridine, 5,6-dihydrouridine, 5-methyl-dihydrouridine (m5D), 2-thio-dihydrouridine, 2-thio-dihydropseudouridine, 2-methoxy-uridine, 2-methoxy- 4-thio-uridine, 4-methoxy-pseudouridine, 4-methoxy-2-thio-pseudouridine, N1-methyl- pseudouridine (also known as 1-methylpseudouridine (m1ψ)), 3-(3-amino-3- carboxypropyl)uridine (acp3U), 1-methyl-3-(3-amino-3-carboxypropyl)pseudouridine (acp3 ψ), 5-(isopentenylaminomethyl)uridine (inm5U), 5-(isopentenylaminomethyl)-2-thio-uridine (inm5s2U), α-thio-uridine, 2′-O-methyl-uridine (Um), 5,2′-O-dimethyl-uridine (m5Um), 2′-O- methyl-pseudouridine (ψm), 2-thio-2′-O-methyl-uridine (s2Um), 5-methoxycarbonylmethyl- 2′-O-methyl-uridine (mcm5Um), 5-carbamoylmethyl-2′-O-methyl-uridine (ncm5Um), 5- carboxymethylaminomethyl-2′-O-methyl-uridine (cmnm5Um), 3,2′-O-dimethyl-uridine (m3Um), 5-(isopentenylaminomethyl)-2′-O-methyl-uridine (inm5Um), 1-thio-uridine, deoxythymidine, 2'‐F‐ara‐uridine, 2'‐F‐uridine, 2'‐OH‐ara‐uridine, 5‐(2‐carbomethoxyvinyl) uridine, and 5‐[3‐(1‐E‐propenylamino)uridine. [0110] In some embodiments, the modified nucleobase is a modified cytosine. Exemplary nucleobases and nucleosides having a modified cytosine include 5-aza-cytidine, 6-aza- cytidine, pseudoisocytidine, 3-methyl-cytidine (m3C), N4-acetyl-cytidine (ac4C), 5-formyl- cytidine (f5C), N4-methyl-cytidine (m4C), 5-methyl-cytidine (m5C), 5-halo-cytidine (e.g., 5- iodo-cytidine), 5-hydroxymethyl-cytidine (hm5C), 1-methyl-pseudoisocytidine, pyrrolo- cytidine, pyrrolo-pseudoisocytidine, 2-thio-cytidine (s2C), 2-thio-5-methyl-cytidine, 4-thio- pseudoisocytidine, 4-thio-1-methyl-pseudoisocytidine, 4-thio-1-methyl-1-deaza- pseudoisocytidine, 1-methyl-1-deaza-pseudoisocytidine, zebularine, 5-aza-zebularine, 5- methyl-zebularine, 5-aza-2-thio-zebularine, 2-thio-zebularine, 2-methoxy-cytidine, 2- methoxy-5-methyl-cytidine, 4-methoxy-pseudoisocytidine, 4-methoxy-1-methyl- pseudoisocytidine, lysidine (k2C), α-thio-cytidine, 2′-O-methyl-cytidine (Cm), 5,2′-O- dimethyl-cytidine (m5Cm), N4-acetyl-2′-O-methyl-cytidine (ac4Cm), N4,2′-O-dimethyl- cytidine (m4Cm), 5-formyl-2′-O-methyl-cytidine (f5Cm), N4,N4,2′-O-trimethyl-cytidine (m4 2Cm), 1-thio-cytidine, 2'‐F‐ara‐cytidine, 2'‐F‐cytidine, and 2'‐OH‐ara‐cytidine. [0111] In some embodiments, the modified nucleobase is a modified adenine. Exemplary nucleobases and nucleosides having a modified adenine include 2-amino-purine, 2, 6- diaminopurine, 2-amino-6-halo-purine (e.g., 2-amino-6-chloro-purine), 6-halo-purine (e.g., 6- chloro-purine), 2-amino-6-methyl-purine, 8-azido-adenosine, 7-deaza-adenine, 7-deaza-8- aza-adenine, 7-deaza-2-amino-purine, 7-deaza-8-aza-2-amino-purine, 7-deaza-2,6- diaminopurine, 7-deaza-8-aza-2,6-diaminopurine, 1-methyl-adenosine (m1A), 2-methyl- adenine (m2A), N6-methyl-adenosine (m6A), 2-methylthio-N6-methyl-adenosine (ms2m6A), N6-isopentenyl-adenosine (i6A), 2-methylthio-N6-isopentenyl-adenosine (ms2i6A), N6-(cis- hydroxyisopentenyl)adenosine (io6A), 2-methylthio-N6-(cis-hydroxyisopentenyl)adenosine (ms2io6A), N6-glycinylcarbamoyl-adenosine (g6A), N6-threonylcarbamoyl-adenosine (t6A), N6-methyl-N6-threonylcarbamoyl-adenosine (m6t6A), 2-methylthio-N6-threonylcarbamoyl- adenosine (ms2g6A), N6,N6-dimethyl-adenosine (m6 2A), N6-hydroxynorvalylcarbamoyl- adenosine (hn6A), 2-methylthio-N6-hydroxynorvalylcarbamoyl-adenosine (ms2hn6A), N6- acetyl-adenosine (ac6A), 7-methyl-adenine, 2-methylthio-adenine, 2-methoxy-adenine, α- thio-adenosine, 2′-O-methyl-adenosine (Am), N6,2′-O-dimethyl-adenosine (m6Am), N6,N6,2′-O-trimethyl-adenosine (m62Am), 1,2′-O-dimethyl-adenosine (m1Am), 2′-O- ribosyladenosine (phosphate) (Ar(p)), 2-amino-N6-methyl-purine, 1-thio-adenosine, 8-azido- adenosine, 2'‐F‐ara‐adenosine, 2'‐F‐adenosine, 2'‐OH‐ara‐adenosine, and N6‐(19‐amino‐ pentaoxanonadecyl)-adenosine. [0112] In some embodiments, the modified nucleobase is a modified guanine. Exemplary nucleobases and nucleosides having a modified guanine include inosine (I), 1-methyl-inosine (m1I), wyosine (imG), methylwyosine (mimG), 4-demethyl-wyosine (imG-14), isowyosine (imG2), wybutosine (yW), peroxywybutosine (o2yW), hydroxywybutosine (OHyW), undermodified hydroxywybutosine (OHyW*), 7-deaza-guanosine, queuosine (Q), epoxyqueuosine (oQ), galactosyl-queuosine (galQ), mannosyl-queuosine (manQ), 7-cyano-7- deaza-guanosine (preQ0), 7-aminomethyl-7-deaza-guanosine (preQ1), archaeosine (G+), 7- deaza-8-aza-guanosine, 6-thio-guanosine, 6-thio-7-deaza-guanosine, 6-thio-7-deaza-8-aza- guanosine, 7-methyl-guanosine (m7G), 6-thio-7-methyl-guanosine, 7-methyl-inosine, 6- methoxy-guanosine, 1-methyl-guanosine (m1G), N2-methyl-guanosine (m2G), N2,N2- dimethyl-guanosine (m22G), N2,7-dimethyl-guanosine (m2,7G), N2, N2,7-dimethyl-guanosine (m2,2,7G), 8-oxo-guanosine, 7-methyl-8-oxo-guanosine, 1-methyl-6-thio-guanosine, N2- methyl-6-thio-guanosine, N2,N2-dimethyl-6-thio-guanosine, α-thio-guanosine, 2′-O-methyl- guanosine (Gm), N2-methyl-2′-O-methyl-guanosine (m2Gm), N2,N2-dimethyl-2′-O-methyl- guanosine (m2 2Gm), 1-methyl-2′-O-methyl-guanosine (m1Gm), N2,7-dimethyl-2′-O-methyl- guanosine (m2,7Gm), 2′-O-methyl-inosine (Im), 1,2′-O-dimethyl-inosine (m1Im), and 2′-O- ribosylguanosine (phosphate) (Gr(p)). [0113] The nucleobase of the nucleotide can be independently selected from a purine, a pyrimidine, a purine or pyrimidine analog. For example, the nucleobase can each be independently selected from adenine, cytosine, guanine, uracil, or hypoxanthine. In another embodiment, the nucleobase can also include, for example, naturally-occurring and synthetic derivatives of a base, including pyrazolo[3,4-d]pyrimidines, 5-methylcytosine (5-me-C), 5- hydroxymethyl cytosine, xanthine, hypoxanthine, 2-aminoadenine, 6-methyl and other alkyl derivatives of adenine and guanine, 2-propyl and other alkyl derivatives of adenine and guanine, 2-thiouracil, 2-thiothymine and 2-thiocytosine, 5-propynyl uracil and cytosine, 6- azo uracil, cytosine and thymine, 5-uracil (pseudouracil), 4-thiouracil, 8-halo (e.g., 8-bromo), 8-amino, 8-thiol, 8-thioalkyl, 8-hydroxyl and other 8-substituted adenines and guanines, 5- halo particularly 5-bromo, 5-trifluoromethyl and other 5-substituted uracils and cytosines, 7- methylguanine and 7-methyladenine, 8-azaguanine and 8-azaadenine, deazaguanine, 7- deazaguanine, 3-deazaguanine, deazaadenine, 7-deazaadenine, 3-deazaadenine, pyrazolo[3,4- d]pyrimidine, imidazo[1,5-a]1,3,5 triazinones, 9-deazapurines, imidazo[4,5-d]pyrazines, thiazolo[4,5-d]pyrimidines, pyrazin-2-ones, 1,2,4-triazine, pyridazine; and 1,3,5 triazine. [0114] Particular embodiments of the nucleic acid-based compositions described herein comprise modifications including 5-methylcytosine (“5-me-C”) bases or 5-methylcytidine (m5c) nucleotides. [0115] The nucleic acid-based compositions of the present disclosure may or may not be uniformly modified along the entire length of the molecule. For example, one or more or all types of nucleotide (e.g., purine or pyrimidine, or any one or more or all of A, G, T/U, C) may or may not be uniformly modified in the nucleic acid-based compositions described herein. In some embodiments, all nucleotides X in a nucleic acid-based composition are modified, wherein X may be any one of nucleotides A, G, T/U, C, or any one of the combinations A+G, A+T/U, A+C, G+T/U, G+C, T/U+C, A+G+T/U, A+G+C, G+T/U+C or A+G+C. [0116] Different sugar modifications, nucleotide modifications, and/or internucleoside linkages (e.g., backbone structures) may be introduced at various positions in a nucleic acid- based composition. One of ordinary skill in the art will appreciate that the nucleotide analogs or other modification(s) may be located at any position(s) of a nucleic acid-based composition such that the function of the nucleic acid-based composition is not substantially decreased. The nucleic acid-based composition of the present disclosure may contain from about 1% to about 100% modified nucleotides (either in relation to overall nucleotide content, or in relation to one or more types of nucleotide, i.e. any one or more of A, G, T/U or C) or any intervening percentage (e.g., from 1% to 20%, from 1% to 25%, from 1% to 50%, from 1% to 60%, from 1% to 70%, from 1% to 80%, from 1% to 90%, from 1% to 95%, from 10% to 20%, from 10% to 25%, from 10% to 50%, from 10% to 60%, from 10% to 70%, from 10% to 80%, from 10% to 90%, from 10% to 95%, from 10% to 100%, from 20% to 25%, from 20% to 50%, from 20% to 60%, from 20% to 70%, from 20% to 80%, from 20% to 90%, from 20% to 95%, from 20% to 100%, from 50% to 60%, from 50% to 70%, from 50% to 80%, from 50% to 90%, from 50% to 95%, from 50% to 100%, from 70% to 80%, from 70% to 90%, from 70% to 95%, from 70% to 100%, from 80% to 90%, from 80% to 95%, from 80% to 100%, from 90% to 95%, from 90% to 100%, and from 95% to 100%). [0117] In some embodiments, the nucleic acid-based compositions of the present disclosure may be modified to be a circular nucleic acid. The termini of the nucleic acid- based compositions of the present disclosure may be linked by chemical reagents or enzymes, producing circular nucleic acid-based compositions that have no free ends. Circular nucleic acid-based compositions are expected to be more stable than linear counterparts and to be resistant to digestion with exonucleases. Circular nucleic acid-based compositions may further comprise other structural and/or chemical modifications with respect to A, G, T/U or C ribonucleotides/deoxyribonucleotides. [0118] The nucleic acid-based compositions of the present disclosure may comprise a combination of modifications. The nucleic acid-based compositions may comprise at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30 modifications or modified nucleotides. [0119] In some embodiments, the nucleic acid-based compositions are at least 50% modified, e.g., at least 50% of the nucleotides are modified. In some embodiments, the nucleic acid-based compositions are at least 75% modified, e.g., at least 75% of the nucleotides are modified. It is to be understood that since a nucleotide (sugar, base and phosphate moiety, e.g., linker) may each be modified, any modification to any portion of a nucleotide, or nucleoside, will constitute a modification. [0120] In some embodiments, the nucleic acid-based compositions are at least 10% modified in only one component of the nucleotide, with such component being the nucleobase, sugar, or linkage between nucleosides. For example, modifications may be made to at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 100% of the nucleobases, sugars, or linkages of a nucleic acid-based composition described herein. Sugar Modifications [0121] In some embodiments, a nucleic acid-based composition comprises at least one sugar modification. Generally, RNA includes the sugar group ribose, which is a 5-membered ring having an oxygen. Exemplary, non-limiting modified nucleotides include replacement of the oxygen in ribose (e.g., with S, Se, or alkylene, such as methylene or ethylene); addition of a double bond (e.g., to replace ribose with cyclopentenyl or cyclohexenyl); ring contraction of ribose (e.g., to form a 4-membered ring of cyclobutane or oxetane); ring expansion of ribose (e.g., to form a 6- or 7-membered ring having an additional carbon or heteroatom, such as for anhydrohexitol, altritol, mannitol, cyclohexanyl, cyclohexenyl, and morpholino that also has a phosphoramidate backbone); multicyclic forms (e.g., tricyclo; and “unlocked” forms, such as glycol nucleic acid (GNA) (e.g., R-GNA or S-GNA, where ribose is replaced by glycol units attached to phosphodiester bonds), threose nucleic acid (TNA, where ribose is replace with α-L-threofuranosyl-(3′→2′)) , and peptide nucleic acid (PNA, where 2-amino-ethyl-glycine linkages replace the ribose and phosphodiester backbone). The sugar group can also contain one or more carbons that possess the opposite stereochemical configuration than that of the corresponding carbon in ribose. Thus, a nucleic acid-based composition can include nucleotides containing, e.g., arabinose, as the sugar. [0122] Nonlimiting examples of the sugar modification may include the modifications provided in Table 10. The nucleic acid-based compositions of the present disclosure can have one or more nucleotides carrying a modification as provided in Table 10. In some embodiments, each of the nucleotides of a nucleic acid-based composition as described herein carries any one of the modifications as provided in Table 10, or none of the modifications as provided in Table 10. Table 10. Nucleotide Sugar Modifications
Figure imgf000166_0001
Figure imgf000167_0001
Figure imgf000168_0001
Figure imgf000169_0001
Figure imgf000170_0001
[0123] In some embodiments, at least one of the 2' positions of the sugar (OH in RNA or H in DNA) of a nucleotide of the nucleic acid-based composition is substituted with -OMe, referred to as 2’-OMe. In some embodiments, at least one of the 2' positions of the sugar (OH in RNA or H in DNA) of a nucleotide of the nucleic acid-based composition is substituted with -F, referred to as 2’-F. Internucleoside Linkages and Other Modifications [0124] The nucleic acid-based compositions of the present disclosure can include any modification to the internucleoside linkage (e.g. to a linking phosphate / to a phosphodiester linkage / to the phosphodiester backbone). Examples of modified phosphate groups include, but are not limited to, phosphorothioate, methylphosphonate, phosphoroselenates, boranophosphates, boranophosphate esters, hydrogen phosphonates, phosphoramidates, phosphorodiamidates, alkyl or aryl phosphonates, and phosphotriesters. Phosphorodithioates have both non-linking oxygens replaced by sulfur. The phosphate linker can also be modified by the replacement of a linking oxygen with nitrogen (bridged phosphoramidates), sulfur (bridged phosphorothioates), and carbon (bridged methylene-phosphonates). [0125] In some embodiments, the nucleic acid-based composition comprises at least one phosphorothioate linkage or methylphosphonate linkage between nucleotides. [0126] In some embodiments, the nucleic acid-based composition comprises 3’ and/or 5’ capping or overhang. In some embodiments, the nucleic acid-based composition of the present disclosure may comprise at least one inverted deoxyribonucleoside overhang (e.g., dT). The inverted overhang, e.g., dT, may be at the 5’ terminus or 3’ terminus of the nucleic acid-based composition. In some embodiments, the nucleic acid-based composition of the present disclosure comprises inverted abasic modifications. The inverted abasic modification(s) can be on the 5’ end, or 3’ end, or both ends of the nucleic acid-based composition. [0127] In some embodiments, the nucleic acid-based composition comprises at least one 5’-(E)-vinylphosphonate (5’-E-VP), a phosphate mimic, as a modification. [0128] In some embodiments, the nucleic acid-based composition comprises at least one glycol nucleic acid (GNA), an acyclic nucleic acid analogue, as a modification. [0129] In a preferred embodiment, the GABP1-hybridizing nucleic acid-based compositions described herein are 14-mers. As an example, a nucleic acid-based composition described herein can be a 14-mer, i.e., having 14 nucleotides. The 14-mers can be fully modified or partially modified according to the modifications provided herein. Configurations of varying 14-mer nucleic acid-based compositions with varying modifications can be represented as follows. For example, a 2-10-2 ASO is a 14-mer ASO having a configuration wherein two 5’ nucleotides carry a first defined modification, 10 internal nucleotides have a second defined modification, and two 3’ nucleotides carry the first defined modification, or a third modification. The 14-mer ASOs described herein can have a 1-12-1 configuration, a 2-10-2 configuration, a 3-8-3 configuration, and so on. Alternatively, the 14-mer ASOs described herein can have a 1-13 configuration, a 2-12 configuration, a 3- 11 configuration, and so on. Likewise, the 14-mer ASOs can have a 13-1 configuration, a 12- 2 configuration, a 11-3 configuration, and so on. In some embodiments, the 14-mer ASOs can have a 1-11-2 configuration, a 1-10-3 configuration, a 1-9-4 configuration, and so on. Similarly, the 14-mer ASOs can have a 2-11-1 configuration, a 3-10-1 configuration, a 4-9-1 configuration, and so on. In some embodiments, GABPB1-hybridizing oligonucleotides are 2-10-2 LNA-2’MOE-LNA fully modified 14-mer ASOs, wherein the two 5’ nucleotides are locked nucleic acids, the ten internal nucleotides carry 2‘-O-(2-Methoxyethyl) modifications, and the two 3’ nucleotides are locked nucleic acids. [0130] In another preferred embodiment, the GABPB1-hybridizing nucleic acid-based compositions described herein are 15-mers. As an example, a nucleic acid-based composition described herein can be a 15-mer, i.e., having 15 nucleotides. The 15-mers can be fully modified or partially modified according to the modifications provided herein. Configurations of varying 15-mer nucleic acid-based compositions with varying modifications can be represented as follows. For example, a 2-11-2 ASO is a 15-mer ASO having a configuration wherein two 5’ nucleotides carry a first defined modification, 11 internal nucleotides have a second defined modification, and two 3’ nucleotides carry the first defined modification, or a third modification. The 15-mer ASOs described herein can have a 1-13-1 configuration, a 2-11-2 configuration, a 3-9-3 configuration, and so on. Alternatively, the 15-mer ASOs described herein can have a 1-14 configuration, a 2-13 configuration, a 3- 12 configuration, and so on. Likewise, the 15-mer ASOs can have a 14-1 configuration, a 13- 2 configuration, a 12-3 configuration, and so on. In some embodiments, the 15-mer ASOs can have a 1-12-2 configuration, a 1-11-3 configuration, a 1-10-4 configuration, and so on. Similarly, the 15-mer ASOs can have a 2-12-1 configuration, a 3-11-1 configuration, a 4-10-1 configuration, and so on. In some embodiments, GABPB1-hybridizing oligonucleotides are 2-11-2 LNA-2’MOE-LNA fully modified 15-mer ASOs, wherein the two 5’ nucleotides are locked nucleic acids, the eleven internal nucleotides carry 2‘-O-(2-Methoxyethyl) modifications, and the two 3’ nucleotides are locked nucleic acids. [0131] In another preferred embodiment, the GABPB1-hybridizing nucleic acid-based compositions described herein are 16-mers. As an example, a nucleic acid-based composition described herein can be a 16-mer, i.e., having 16 nucleotides. The 16-mers can be fully modified or partially modified according to the modifications provided herein. Configurations of varying 16-mer nucleic acid-based compositions with varying modifications can be represented as follows. For example, a 2-12-2 ASO is a 16-mer ASO having a configuration wherein two 5’ nucleotides carry a first defined modification, 12 internal nucleotides have a second defined modification, and two 3’ nucleotides carry the first defined modification, or a third modification. The 16-mer ASOs described herein can have a 1-14-1 configuration, a 2-12-2 configuration, a 3-10-3 configuration, and so on. Alternatively, the 16-mer ASOs described herein can have a 1-15 configuration, a 2-14 configuration, a 3-13 configuration, and so on. Likewise, the 16-mer ASOs can have a 15-1 configuration, a 14-2 configuration, a 13-3 configuration, and so on. In some embodiments, the 16-mer ASOs can have a 1-13-2 configuration, a 1-12-3 configuration, a 1-11-4 configuration, and so on. Similarly, the 16-mer ASOs can have a 2-13-1 configuration, a 3- 12-1 configuration, a 4-11-1 configuration, and so on. In some embodiments, GABPB1- hybridizing oligonucleotides are 2-10-2 LNA-2’MOE-LNA fully modified 16-mer ASOs, wherein the two 5’ nucleotides are locked nucleic acids, the twelve internal nucleotides carry 2‘-O-(2-Methoxyethyl) modifications, and the two 3’ nucleotides are locked nucleic acids. [0132] In another preferred embodiment, the GABPB1-hybridizing nucleic acid-based compositions described herein are 20-mers. As an example, a nucleic acid-based composition described herein can be a 20-mer, i.e., having 20 nucleotides. The 20-mers can be fully modified or partially modified according to the modifications provided herein. Configurations of varying 20-mer nucleic acid-based compositions with varying modifications can be represented as follows. For example, a 5-10-5 ASO is a 20-mer ASO having a configuration wherein five 5’ nucleotides carry a first defined modification, 10 internal nucleotides have a second defined modification, and five 3’ nucleotides carry the first defined modification, or a third modification. The 20-mer ASOs described herein can have a 1-18-1 configuration, a 2-16-2 configuration, a 3-14-3 configuration, and so on. Alternatively, the 20-mer ASOs described herein can have a 1-19 configuration, a 2-18 configuration, a 3-17 configuration, and so on. Likewise, the 20-mer ASOs can have a 19-1 configuration, a 18-2 configuration, a 17-3 configuration, and so on. In some embodiments, the 20-mer ASOs can have a 1-17-2 configuration, a 1-16-3 configuration, a 1-5-4 configuration, and so on. Similarly, the 20-mer ASOs can have a 2-17-1 configuration, a 3- 16-1 configuration, a 4-15-1 configuration, and so on. In some embodiments, GABP- hybridizing oligonucleotides are 5-10-5 LNA-2’MOE-LNA fully modified 20-mer ASOs, wherein the five 5’ nucleotides are locked nucleic acids, the ten internal nucleotides carry 2‘- O-(2-Methoxyethyl) modifications, and the five 3’ nucleotides are locked nucleic acids. Conjugates and Combinations [0133] Conjugation may result in increased stability and/or half-life and may be particularly useful in targeting the nucleic acid-based compositions of the present disclosure to specific sites in the cell, tissue, or organism. The nucleic acid-based compositions of the present disclosure can be designed to be conjugated to other polynucleotides, dyes, intercalating agents (e.g. acridines), cross-linkers (e.g. psoralene, mitomycin C), porphyrins (TPPC4, texaphyrin, Sapphyrin), polycyclic aromatic hydrocarbons (e.g., phenazine, dihydrophenazine), artificial endonucleases (e.g. EDTA), alkylating agents, phosphate, amino, mercapto, PEG (e.g., PEG-40K), MPEG, [MPEG]2, polyamino, alkyl, substituted alkyl, radiolabeled markers, enzymes, haptens (e.g. biotin), transport/absorption facilitators (e.g., aspirin, vitamin E, folic acid), synthetic ribonucleases, proteins, e.g., glycoproteins, or peptides, e.g., molecules having a specific affinity for a co-ligand, or antibodies e.g., an antibody, that binds to a specified cell type such as a kidney tubule cell, cancer cell, endothelial cell, or bone cell, hormones and hormone receptors, non-peptidic species, such as lipids, lectins, carbohydrates, vitamins, cofactors, or a drug. Suitable conjugates for nucleic acid molecules are disclosed in International Publication WO 2013/090648, the contents of which are incorporated herein by reference in their entirety. [0134] According to the present disclosure, nucleic acid-based compositions can be administered with, or further comprise one or more of: RNAi agents, small interfering RNAs (siRNAs), small hairpin RNAs (shRNAs), long non-coding RNAs (lncRNAs), enhancer RNAs, enhancer-derived RNAs or enhancer-driven RNAs (eRNAs), microRNAs (miRNAs), miRNA binding sites, antisense RNAs, ribozymes, catalytic DNA, tRNA, RNAs that induce triple helix formation, aptamers or vectors, and the like to achieve different functions. The one or more RNAi agents, small interfering RNAs (siRNAs), small hairpin RNAs (shRNAs), long non-coding RNAs (lncRNA), microRNAs (miRNAs), miRNA binding sites, antisense RNAs, ribozymes, catalytic DNA, tRNA, RNAs that induce triple helix formation, aptamers or vectors may comprise at least one modification or substitution. [0135] In some embodiments, the modification is a chemical substitution of the nucleic acid at a sugar position, a chemical substitution at a phosphate position, or a chemical substitution at a base position. In some embodiments, the chemical modification is incorporation of a modified nucleotide; 3′ capping; conjugation to a high molecular weight, non-immunogenic compound; conjugation to a lipophilic compound; or incorporation of phosphorothioate into the phosphate backbone. In some embodiments, the high molecular weight, non-immunogenic compound is polyalkylene glycol, or polyethylene glycol (PEG). [0136] In some embodiments, nucleic acid-based compositions of the present disclosure may be attached to a transgene so as to be co-expressed from an RNA polymerase II promoter. In a non-limiting example, nucleic acid-based compositions are attached to green fluorescent protein gene (GFP) or other reporter or tag. [0137] In some embodiments, nucleic acid-based compositions of the present disclosure may be attached to a DNA or RNA aptamer, thereby producing a nucleic acid-based composition-aptamer conjugate. Aptamers are oligonucleotides or peptides with high selectivity, affinity, and stability. They assume specific and stable three-dimensional shapes, thereby providing highly specific, tight binding to target molecules. An aptamer may be a nucleic acid species that has been engineered through repeated rounds of in vitro selection or equivalently, SELEX (systematic evolution of ligands by exponential enrichment) to bind to various molecular targets such as small molecules, proteins, nucleic acids, and even cells, tissues and organisms. Nucleic acid aptamers have specific binding affinity to molecules through interactions other than classic Watson-Crick base pairing. Nucleic acid aptamers, like peptides generated by phage display or monoclonal antibodies (mAbs), are capable of specifically binding to selected targets and, through binding, block their targets’ ability to function. In some cases, aptamers may also be peptide aptamers. For any specific molecular target, nucleic acid aptamers can be identified from combinatorial libraries of nucleic acids, e.g. by SELEX. Peptide aptamers may be identified using a yeast two hybrid system. A skilled person is therefore able to design suitable aptamers for delivering the nucleic acid- based compositions of the present disclosure to target cells such as kidney cells or specific sub-populations of kidney cells, like tubule cells. DNA aptamers, RNA aptamers and peptide aptamers are contemplated. Administration of nucleic acid-based compositions including ASOs of the present disclosure to the kidney using kidney-specific aptamers is specifically contemplated. [0138] As used herein, a typical nucleic acid aptamer is approximately 10-15 kDa in size (20-45 nucleotides), binds its target with at least nanomolar affinity, and discriminates against closely related targets. Nucleic acid aptamers may be ribonucleic acid, deoxyribonucleic acid, or mixed ribonucleic acid and deoxyribonucleic acid. Aptamers may be single-stranded ribonucleic acid, deoxyribonucleic acid or mixed ribonucleic acid and deoxyribonucleic acid. Aptamers may comprise at least one chemical modification. [0139] A suitable nucleotide length for an aptamer ranges from about 15 to about 100 nucleotides (nt), 15-30 nt, 20-25 nt, 30-100 nt, 30-60 nt, 25-70 nt, 25-60 nt, 40-60 nt, 25-40 nt, 30-40 nt, any of 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39 or 40 nt or 40-70 nt in length. However, the sequence can be designed with sufficient flexibility such that it can accommodate interactions of aptamers with two targets at the distances described herein. Aptamers may be further modified to provide protection from nuclease and other enzymatic activities. The aptamer sequence can be modified by any suitable methods known in the art. [0140] Nucleic acid-based composition-aptamer conjugates may be formed using any known method for linking two moieties, such as direct chemical bond formation or linkage via a linker such as streptavidin. [0141] In some embodiments, nucleic acid-based compositions of the present disclosure may be attached to an antibody. Methods of generating antibodies against a target cell surface receptor are well known. The nucleic acid-based compositions may be attached to such antibodies with known methods, for example using RNA carrier proteins. The resulting complex may then be administered to a subject and taken up by the target cells via receptor- mediated endocytosis. [0142] In some embodiments, nucleic acid-based compositions of the present disclosure may be conjugated with lipid moieties such as a cholesterol moiety (Letsinger et al., Proc. Natl. Acid. Sci. USA, 1989, 86: 6553-6556), cholic acid (Manoharan et al., Biorg. Med. Chem. Let., 1994, 4:1053-1060), a thioether, e.g., beryl-5-tritylthiol (Manoharan et al., Ann. N.Y. Acad. Sci., 1992, 660:306-309; Manoharan et al., Biorg. Med. Chem. Let., 1993, 3:2765-2770), a thiocholesterol (Oberhauser et al., Nucl. Acids Res., 1992, 20:533-538), an aliphatic chain, e.g., dodecandiol or undecyl residues (Saison-Behmoaras et al., EMBO J, 1991, 10:1111-1118; Kabanov et al., FEBS Lett., 1990, 259:327-330; Svinarchuk et al., Biochimie, 1993, 75:49-54), a phospholipid, e.g., di-hexadecyl-rac-glycerol or triethyl- ammonium 1,2-di-O-hexadecyl-rac-glycero-3-Hphosphonate (Manoharan et al., Tetrahedron Lett., 1995, 36:3651-3654; Shea et al., Nucl. 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 (Manoharan et al., Tetrahedron Lett., 1995, 36:3651-3654), a palmityl moiety (Mishra et al., Biochim. Biophys. Acta, 1995, 1264:229-237), or an octadecylamine or hexylamino-carbonyloxycholesterol moiety (Crooke et al., J. Pharmacol. Exp. Ther., 1996, 277:923-937), the content of each of which is herein incorporated by reference in its entirety. [0143] In some embodiments, the nucleic acid-based compositions of the present disclosure are conjugated with a ligand. In a non-limiting example, the ligand may be any ligand disclosed in US 20130184328 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety. The conjugate has a formula of Ligand- [linker]optional-[tether]optional-oligonucleotide agent. The oligonucleotide agent may comprise a subunit having formula (I) disclosed by US 20130184328. In another non-limiting example, the ligand may be any ligand disclosed in US 20130317081 to Akinc et al., the contents of which are incorporated herein by reference in their entirety, such as a lipid, a protein, a hormone, or a carbohydrate ligand of Formula II-XXVI. The ligand may be coupled with the nucleic acid-based composition with a bivalent or trivalent branched linker in Formula XXXI-XXXV disclosed in Akinc. [0144] Representative U.S. patents that teach the preparation of such nucleic acid/lipid conjugates include, but are not limited to, U.S. Pat. Nos. 4,828,979; 4,948,882; 5,218,105; 5,525,465; 5,541,313; 5,545,730; 5,552,538; 5,578,717, 5,580,731; 5,591,584; 5,109,124; 5,118,802; 5,138,045; 5,414,077; 5,486,603; 5,512,439; 5,578,718; 5,608,046; 4,587,044; 4,605,735; 4,667,025; 4,762,779; 4,789,737; 4,824,941; 4,835,263; 4,876,335; 4,904,582; 4,958,013; 5,082,830; 5,112,963; 5,214,136; 5,082,830; 5,112,963; 5,214,136; 5,245,022; 5,254,469; 5,258,506; 5,262,536; 5,272,250; 5,292,873; 5,317,098; 5,371,241, 5,391,723; 5,416,203, 5,451,463; 5,510,475; 5,512,667; 5,514,785; 5,565,552; 5,567,810; 5,574,142; 5,585,481; 5,587,371; 5,595,726; 5,597,696; 5,599,923; 5,599,928 and 5,688,941, the content of each of which is herein incorporated by reference in its entirety. [0145] The nucleic acid-based compositions of the present disclosure may be provided in combination with other active ingredients known to have an effect in the particular method being considered. The other active ingredients may be administered simultaneously, separately, or sequentially with the nucleic acid-based compositions. In some embodiments, nucleic acid-based compositions of the present disclosure are administered with nucleic acid- based compositions, ASOs, or other agents modulating a different target gene. [0146] In some embodiments, the nucleic acid-based compositions are conjugated with a carbohydrate ligand, such as any carbohydrate ligand disclosed in US Pat No. 8106022 and 8828956 to Manoharan et al. (Alnylam Pharmaceuticals), the contents of which are incorporated herein by reference in their entireties. For example, the carbohydrate ligand may be monosaccharide, disaccharide, trisaccharide, tetrasaccharide, oligosaccharide, or polysaccharide. These carbohydrate-conjugated RNA agents may target a particular cell or tissue type. In some embodiments, the nucleic acid-based compositions are conjugated with more than one carbohydrate ligand, preferably two or three. In some embodiments, the nucleic acid-based compositions are conjugated with one or more galactose moiety. In some embodiments, the nucleic acid-based compositions are conjugated with at least one (e.g., two or three or more) lactose molecules (lactose is a glucose coupled to a galactose). In some embodiments, the nucleic acid-based compositions are conjugated with at least one (e.g., two or three or more) N-Acetyl-Galactosamine (GalNAc), N-Ac-Glucosamine (GluNAc), or mannose (e.g., mannose-6-phosphate). [0147] In some embodiments, nucleic acid-based compositions of the present disclosure are administered with a small interfering RNA or siRNA that inhibits the expression of a gene. [0148] In some embodiments, nucleic acid-based compositions of the present disclosure are administered with one or more drugs for therapeutic purposes, including for treatment of ADPKD. II. Compositions [0149] One aspect of the present disclosure provides pharmaceutical compositions comprising at least one pharmaceutically acceptable carrier and a nucleic acid-based composition, i.e., oligo- or polynucleotide molecule, including in some instances an antisense oligonucleotide (ASO) that downregulates a target gene. Expression Systems [0150] In some embodiments, antisense oligonucleotides (ASOs) described herein may be operably linked to one or more regulatory nucleotide sequences in an expression construct. Regulatory nucleotide sequences will generally be appropriate for a host cell used for expression. Numerous types of appropriate expression vectors and suitable regulatory sequences are known in the art for a variety of host cells. Typically, the one or more regulatory nucleotide sequences may include, but are not limited to, promoter sequences, leader or signal sequences, transcriptional start and termination sequences, and enhancer or activator sequences. Constitutive or inducible promoters as known in the art are also contemplated. The promoters may be either naturally occurring promoters, or hybrid promoters that combine elements of more than one promoter. An expression construct may be present in a cell on an episome, such as a plasmid, or the expression construct may be inserted in a chromosome. In a specific embodiment, the expression vector includes a selectable marker gene to allow the selection of transformed host cells. Certain embodiments include an expression vector encoding a GABPB1-hybridizing oligonucleotide sequence operably linked to at least one regulatory sequence. Regulatory sequences for use herein include promoters, enhancers, and other expression control elements. In certain embodiments, an expression vector is designed considering the choice of the host cell to be transformed, the particular GABP1L sequence to be expressed, the vector's copy number, the ability to control that copy number, or the expression of other proteins encoded by the vector, such as antibiotic markers. [0151] In some embodiments, the gene products of the combinatorial libraries generated by the combinatorial mutagenesis of the nucleic acids described herein may be screened. Such screening methods include, for example, cloning the gene library into replicable expression vectors, transforming appropriate cells with the resulting library of vectors, and expressing the combinatorial genes under conditions to form such library. The screening methods optionally further comprise detecting a desired activity and isolating a product detected. Each of the illustrative assays described below are amenable to high-throughput analysis as necessary to screen large numbers of degenerate sequences created by combinatorial mutagenesis techniques. [0152] In some embodiments, the nucleic acids described herein may be expressed in microorganisms. As a non-limiting example, the nucleic acid may be expressed in a bacterial system, for example, in Bacillus brevis, Bacillus megaterium, Bacillus subtilis, Caulobacter crescentus, Escherichia coli and their derivatives. Exemplary promoters include the l- arabinose inducible araBAD promoter (PBAD), the lac promoter, the l-rhamnose inducible rhaP BAD promoter, the T7 RNA polymerase promoter, the trc and tac promoter, the lambda phage promoter Pl, and the anhydrotetracycline-inducible tetA promoter/operator. [0153] In some embodiments, the nucleic acids described herein may be expressed in a yeast expression system. Non-limiting examples of promoters which may be used in yeast vectors include the promoters for 3-phosphoglycerate kinase (Hitzeman et al., J. Biol. Chem. 255:2073 (1980)); other glycolytic enzymes (Hess et al., J. Adv. Enzyme Res. 7:149 (1968); Holland et al., Biochemistry 17:4900 (1978). Others promoters are from, e.g., enolase, glyceraldehyde-3-phosphate dehydrogenase, hexokinase, pyruvate decarboxylase, phosphofructokinase, glucose-6-phosphate isomerase, 3-phosphoglycerate mutase, pyruvate kinase, triosephosphate isomerase, phosphoglucose isomerase, glucokinase alcohol oxidase I (AOX1), alcohol dehydrogenase 2, isocytochrome C, acid phosphatase, degradative enzymes associated with nitrogen metabolism, and the aforementioned glyceraldehyde-3-phosphate dehydrogenase, and enzymes responsible for maltose and galactose utilization. Any plasmid vector containing a yeast-compatible promoter and termination sequences, with or without an origin of replication, is suitable. Certain yeast expression systems are commercially available, for example, from Clontech Laboratories, Inc. (Palo Alto, Calif., e.g. Pyex 4T family of vectors for S. cerevisiae), Invitrogen (Carlsbad, Calif., e.g. Ppicz series Easy Select Pichia Expression Kit) and Stratagene (La Jolla, Calif., e.g. ESP.TM. Yeast Protein Expression and Purification System for S. pombe and Pesc vectors for S. cerevisiae). [0154] In some embodiments, the nucleic acids described herein may be expressed in mammalian expression systems. Non-limiting examples of mammalian promoters include, for example, promoters from the following genes: ubiquitin/S27a promoter of the hamster (WO 97/15664), Simian vacuolating virus 40 (SV40) early promoter, adenovirus major late promoter, mouse metallothionein-I promoter, the long terminal repeat region of Rous Sarcoma Virus (RSV), mouse mammary tumor virus promoter (MMTV), Moloney murine leukemia virus Long Terminal repeat region, and the early promoter of human Cytomegalovirus (CMV). Examples of other heterologous mammalian promoters are the actin, immunoglobulin or heat shock promoter(s). In a specific embodiment, a yeast alcohol oxidase promoter is used. [0155] In some embodiments, promoters for use in mammalian host cells can be obtained from the genomes of viruses such as polyoma virus, fowlpox virus (UK 2,211,504 published 5 Jul. 1989), bovine papilloma virus, avian sarcoma virus, cytomegalovirus, a retrovirus, hepatitis-B virus and Simian Virus 40 (SV40). In further embodiments, heterologous mammalian promoters are used. Examples include the actin promoter, an immunoglobulin promoter, and heat-shock promoters. The early and late promoters of SV40 are conveniently obtained as an SV40 restriction fragment which also contains the SV40 viral origin of replication. Fiers et al., Nature 273: 113-120 (1978). The immediate early promoter of the human cytomegalovirus is conveniently obtained as a HindIII E restriction fragment. Greenaway, P. J. et al., Gene 18: 355-360 (1982). The foregoing references are incorporated by reference in their entirety. [0156] In some embodiments, the nucleic acids described herein may be expressed in insect cell expression systems. Eukaryotic expression systems employing insect cell hosts may rely on either plasmid or baculoviral expression systems. Typical insect host cells are derived from the fall army worm (Spodoptera frugiperda). For expression of a foreign protein these cells are infected with a recombinant form of the baculovirus Autographa californica nuclear polyhedrosis virus which has the gene of interest expressed under the control of the viral polyhedron promoter. Other insects infected by this virus include a cell line known commercially as "High 5" (Invitrogen) which is derived from the cabbage looper (Trichoplusia ni). Another baculovirus sometimes used is the Bombyx mori nuclear polyhedorsis virus which infect the silkworm (Bombyx mori). Numerous baculovirus expression systems are commercially available, for example, from Thermo Fisher (Bac-N- BlueTMk or BAC-TO-BACTM Systems), Clontech (BacPAKTM Baculovirus Expression System), Novagen (Bac Vector SystemTM), or others from Pharmingen or Quantum Biotechnologies. Another insect cell host is the common fruit fly, Drosophila melanogaster, for which a transient or stable plasmid based transfection kit is offered commercially by Thermo Fisher (The DESTM System). [0157] In some embodiments, cells are transformed with vectors that express a nucleic acid described herein. Transformation techniques for inserting new genetic material into eukaryotic cells, including animal and plant cells, are well known. Viral vectors may be used for inserting expression cassettes into host cell genomes. Alternatively, the vectors may be transfected into the host cells. Transfection may be accomplished by methods as described in the art such as, but not limited to, calcium phosphate precipitation, electroporation, optical transfection, protoplast fusion, impalefection, and hydrodynamic delivery. Formulation [0158] Pharmaceutical formulations may additionally comprise a pharmaceutically acceptable excipient, which, as used herein, includes, but is not limited to, any and all solvents, dispersion media, diluents, or other liquid vehicles, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, and the like, as suited to the particular dosage form desired. Various excipients for formulating pharmaceutical compositions and techniques for preparing the composition are known in the art (see Remington: The Science and Practice of Pharmacy, 21st Edition, A. R. Gennaro, Lippincott, Williams & Wilkins, Baltimore, MD, 2006; incorporated herein by reference in its entirety). The use of a conventional excipient medium may be contemplated within the scope of the present disclosure, except insofar as any conventional excipient medium may be incompatible with a substance or its derivatives, such as by producing any undesirable biological effect or otherwise interacting in a deleterious manner with any other component(s) of the pharmaceutical composition. [0159] In some embodiments, compositions are administered to humans, human patients or subjects. For the purposes of the present disclosure, the phrase “active ingredient” generally refers to a nucleic acid-based composition to be delivered as described herein. [0160] Although the descriptions of pharmaceutical compositions provided herein are principally directed to pharmaceutical compositions which are suitable for administration to humans, it will be understood by the skilled artisan that such compositions are generally suitable for administration to any other animal, e.g., to non-human animals, e.g. non-human mammals. Modification of pharmaceutical compositions suitable for administration to humans in order to render the compositions suitable for administration to various animals is well understood, and the ordinarily skilled veterinary pharmacologist can design and/or perform such modification with merely ordinary, if any, experimentation. Subjects to which administration of the pharmaceutical compositions is contemplated include, but are not limited to, humans and/or other primates; mammals, including commercially relevant mammals such as cattle, pigs, horses, sheep, cats, dogs, mice, and/or rats; and/or birds, including commercially relevant birds such as poultry, chickens, ducks, geese, and/or turkeys. [0161] Formulations of the pharmaceutical compositions described herein may be prepared by any method known or hereafter developed in the art of pharmacology. In general, such preparatory methods include the step of bringing the active ingredient into association with an excipient and/or one or more other accessory ingredients, and then, if necessary and/or desirable, dividing, shaping and/or packaging the product into a desired single- or multi-dose unit. [0162] A pharmaceutical composition in accordance with the disclosure may be prepared, packaged, and/or sold in bulk, as a single unit dose, and/or as a plurality of single unit doses. As used herein, a “unit dose” is discrete amount of the pharmaceutical composition comprising a predetermined amount of the active ingredient. The amount of the active ingredient is generally equal to the dosage of the active ingredient which would be administered to a subject and/or a convenient fraction of such a dosage such as, for example, one-half or one-third of such a dosage. [0163] Relative amounts of the active ingredient, the pharmaceutically acceptable excipient, and/or any additional ingredients in a pharmaceutical composition in accordance with the disclosure will vary, depending upon the identity, size, and/or condition of the subject treated and further depending upon the route by which the composition is to be administered. By way of example, the composition may comprise between 0.1% and 100%, e.g., between .5 and 50%, between 1-30%, between 5-80%, at least 80% (w/w) active ingredient. [0164] In some embodiments, the formulations described herein may contain at least one nucleic acid-based composition, e.g., ASO. As a non-limiting example, the formulations may contain 1, 2, 3, 4 or 5 nucleic acid-based compositions with different sequences. In some embodiments, the formulation contains at least three nucleic acid-based compositions with different sequences. In some embodiments, the formulation contains at least five nucleic acid- based compositions with different sequences. [0165] The nucleic acid-based compositions of the present disclosure can be formulated using one or more excipients to: (1) increase stability; (2) increase cell transfection; (3) permit the sustained or delayed release (e.g., from a depot formulation of the nucleic acid- based composition); (4) alter the biodistribution (e.g., target the nucleic acid-based composition to specific tissues or cell types); (5) increase the translation of encoded protein in vivo; and/or (6) alter the release profile of encoded protein in vivo. [0166] In addition to traditional excipients such as any and all solvents, dispersion media, diluents, or other liquid vehicles, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, excipients of the present disclosure can include, without limitation, lipidoids, liposomes, lipid nanoparticles, polymers, lipoplexes, core-shell nanoparticles, peptides, proteins, cells transfected with nucleic acid- based compositions (e.g., for transplantation into a subject), hyaluronidase, nanoparticle mimics and combinations thereof. Accordingly, the formulations of the disclosure can include one or more excipients, each in an amount that together increases the stability of the nucleic acid-based compositions and/or increases cell transfection by the nucleic acid-based compositions. Further, the nucleic acid-based compositions of the present disclosure may be formulated using self-assembled nucleic acid nanoparticles. Pharmaceutically acceptable carriers, excipients, and delivery agents for nucleic acids that may be used in the formulation with the nucleic acid-based compositions of the present disclosure are disclosed in International Publication WO 2013/090648, the contents of which are incorporated herein by reference in their entirety. [0167] In some embodiments, the nucleic acid-based compositions of the present disclosure comprise two single RNA strands that are annealed to form a double-stranded oligonucleotide as the active ingredient. The composition further comprises a salt buffer composed of 50mM Tris-HCl, pH 8.0, 100mM NaCl and 5mM EDTA. [0168] In some embodiments, the nucleic acid-based compositions of the present disclosure may be delivered with dendrimers. Dendrimers are highly branched macromolecules. In some embodiments, the nucleic acid-based compositions are complexed with structurally flexible poly(amidoamine) (PAMAM) dendrimers for targeted in vivo delivery. The complex is called a nucleic acid-based composition-dendrimer complex. Dendrimers have a high degree of molecular uniformity, narrow molecular weight distribution, specific size and shape characteristics, and a highly-functionalized terminal surface. The manufacturing process is a series of repetitive steps starting with a central initiator core. Each subsequent growth step represents a new generation of polymers with a larger molecular diameter and molecular weight, and more reactive surface sites than the preceding generation. [0169] PAMAM dendrimers are efficient nucleotide delivery systems that bear primary amine groups on their surface and also a tertiary amine group inside of the structure. The primary amine group participates in nucleotide binding and promotes their cellular uptake, while the buried tertiary amino groups act as a proton sponge in endosomes and enhance the release of nucleic acid into the cytoplasm. These dendrimers protect the nucleic acid-based composition carried by them from ribonuclease degradation and achieves substantial release of nucleic acid-based composition over an extended period of time via endocytosis for efficient gene targeting. The in vivo efficacy of these nanoparticles have previously been evaluated where biodistribution studies show that the dendrimers preferentially accumulate in peripheral blood mononuclear cells and live with no discernible toxicity (see Zhou et al., Molecular Ther. 2011 Vol. 19, 2228-2238, the contents of which are incorporated herein by reference in their entirety). PAMAM dendrimers may comprise a triethanolamine (TEA) core, a diaminobutane (DAB) core, a cystamine core, a diaminohexane (HEX) core, a diaminododecane (DODE) core, or an ethylenediamine (EDA) core. In some embodiments, PAMAM dendrimers comprise a TEA core or a DAB core. Lipidoids [0170] The synthesis of lipidoids has been extensively described and formulations containing these compounds are particularly suited for delivery of oligonucleotides or nucleic acids (see Mahon et al., Bioconjug Chem. 201021:1448-1454; Schroeder et al., J Intern Med. 2010267:9-21; Akinc et al., Nat Biotechnol. 200826:561-569; Love et al., Proc Natl Acad Sci U S A. 2010107:1864-1869; Siegwart et al., Proc Natl Acad Sci U S A. 2011108:12996- 3001; all of which are incorporated herein in their entireties). [0171] While these lipidoids have been used to effectively deliver double-stranded small interfering RNA molecules in rodents and non-human primates (see Akinc et al., Nat Biotechnol. 200826:561-569; Frank-Kamenetsky et al., Proc Natl Acad Sci U S A. 2008 105:11915-11920; Akinc et al., Mol Ther. 200917:872-879; Love et al., Proc Natl Acad Sci U S A. 2010107:1864-1869; Leuschner et al., Nat Biotechnol. 201129:1005-1010; all of which is incorporated herein in their entirety), the present disclosure contemplates their formulation and use in delivering at least one pharmaceutically acceptable carrier, including ASOs. Complexes, micelles, liposomes or particles can be prepared containing these lipidoids and therefore, can result in an effective delivery of the nucleic acid-based compositions following the injection of a lipidoid formulation via localized and/or systemic routes of administration. Lipidoid complexes of nucleic acid-based compositions can be administered by various means including, but not limited to, intravenous (IV), intramuscular (IM), subcutaneous (SC), intraparenchymal (IPa), intrathecal (IT), or intracerebroventricular (ICV) administration. [0172] In vivo delivery of nucleic acids may be affected by many parameters, including, but not limited to, the formulation composition, nature of particle PEGylation, degree of loading, oligonucleotide to lipid ratio, and biophysical parameters such as, but not limited to, particle size (Akinc et al., Mol Ther. 200917:872-879; the contents of which are herein incorporated by reference in its entirety). As an example, small changes in the anchor chain length of poly(ethylene glycol) (PEG) lipids may result in significant effects on in vivo efficacy. Formulations with the different lipidoids, including, but not limited to penta[3-(1- laurylaminopropionyl)]-triethylenetetramine hydrochloride (TETA–5LAP; aka 98N12-5, see Murugaiah et al., Analytical Biochemistry, 401:61 (2010); the contents of which are herein incorporated by reference in its entirety), C12-200 (including derivatives and variants), and MD1, can be tested for in vivo activity. [0173] The lipidoid referred to herein as “98N12-5” is disclosed by Akinc et al., Mol Ther. 200917:872-879 and the contents of which is incorporated by reference in its entirety. [0174] The lipidoid referred to herein as “C12-200” is disclosed by Love et al., Proc Natl Acad Sci U S A. 2010107:1864-1869 and Liu and Huang, Molecular Therapy. 2010669- 670; the contents of both of which are herein incorporated by reference in their entirety. The lipidoid formulations can include particles comprising either 3 or 4 or more components in addition to the nucleic acid-based compositions. As an example, formulations with certain lipidoids, include, but are not limited to, 98N12-5 and may contain 42% lipidoid, 48% cholesterol and 10% PEG (C14 alkyl chain length). As another example, formulations with certain lipidoids, include, but are not limited to, C12-200 and may contain 50% lipidoid, 10% disteroylphosphatidyl choline, 38.5% cholesterol, and 1.5% PEG-DMG. [0175] In some embodiments, nucleic acid-based compositions formulated with a lipidoid for systemic intravenous administration. For example, a final optimized intravenous formulation using nucleic acid-based compositions and comprising a lipid molar composition of 42% 98N12-5, 48% cholesterol, and 10% PEG-lipid with a final weight ratio of about 7.5 to 1 total lipid to nucleic acid-based compositions and a C14 alkyl chain length on the PEG lipid, with a mean particle size of roughly 50–60 nm, can result in the distribution of the formulation to be greater than 90% to the liver. (see, Akinc et al., Mol Ther. 200917:872- 879; the contents of which are herein incorporated by reference in its entirety). In another example, an intravenous formulation using a C12-200 (see published international application WO2010129709, the contents of which is herein incorporated by reference in their entirety) lipidoid may have a molar ratio of 50/10/38.5/1.5 of C12-200/disteroylphosphatidyl choline/cholesterol/PEG-DMG, with a weight ratio of 7 to 1 total lipid to nucleic acid and a mean particle size of 80 nm may be effective to deliver nucleic acid-based compositions (see, Love et al., Proc Natl Acad Sci U S A. 2010107:1864-1869, the contents of which are herein incorporated by reference in its entirety). [0176] In some embodiments, an MD1 lipidoid-containing formulation may be used to effectively deliver nucleic acid-based compositions to hepatocytes in vivo. The characteristics of optimized lipidoid formulations for intramuscular or subcutaneous routes may vary significantly depending on the target cell type and the ability of formulations to diffuse through the extracellular matrix into the blood stream. While a particle size of less than 150 nm may be desired for effective hepatocyte delivery due to the size of the endothelial fenestrae (see, Akinc et al., Mol Ther. 200917:872-879, the contents of which are herein incorporated by reference in its entirety), use of a lipidoid-formulated nucleic acid-based compositions to deliver the formulation to other cells types including, but not limited to, endothelial cells, myeloid cells, and muscle cells may not be similarly size-limited. [0177] Use of lipidoid formulations to deliver siRNA in vivo to other non-hepatocyte cells such as myeloid cells and endothelium has been reported (see Akinc et al., Nat Biotechnol. 200826:561-569; Leuschner et al., Nat Biotechnol. 201129:1005-1010; Cho et al. Adv. Funct. Mater. 200919:3112-3118; 8th International Judah Folkman Conference, Cambridge, MA October 8-9, 2010; the contents of each of which is herein incorporated by reference in its entirety). Effective delivery to myeloid cells, such as monocytes, lipidoid formulations may have a similar component molar ratio. Different ratios of lipidoids and other components including, but not limited to, disteroylphosphatidyl choline, cholesterol and PEG-DMG, may be used to optimize the formulation of nucleic acid-based compositions for delivery to different cell types including, but not limited to, hepatocytes, myeloid cells, muscle cells, etc. For example, the component molar ratio may include, but is not limited to, 50% C12-200, 10% disteroylphosphatidyl choline, 38.5% cholesterol, and %1.5 PEG-DMG (see Leuschner et al., Nat Biotechnol 201129:1005-1010; the contents of which are herein incorporated by reference in its entirety). The use of lipidoid formulations for the localized delivery of nucleic acids to cells via either subcutaneous or intramuscular delivery, may not require all of the formulation components desired for systemic delivery, and as such may comprise only the lipidoid and nucleic acid-based compositions. Liposomes, Lipoplexes, and Lipid Nanoparticles [0178] The nucleic acid-based compositions of the disclosure can be formulated using one or more liposomes, lipoplexes, or lipid nanoparticles. In some embodiments, pharmaceutical compositions of nucleic acid-based compositions include liposomes. Liposomes are artificially-prepared vesicles which may primarily be composed of a lipid bilayer and may be used as a delivery vehicle for the administration of nutrients and pharmaceutical formulations. Liposomes can be of different sizes such as, but not limited to, a multilamellar vesicle (MLV) which may be hundreds of nanometers in diameter and may contain a series of concentric bilayers separated by narrow aqueous compartments, a small unicellular vesicle (SUV) which may be smaller than 50 nm in diameter, and a large unilamellar vesicle (LUV) which may be between 50 and 500 nm in diameter. Liposome design may include, but is not limited to, opsonins or ligands in order to improve the attachment of liposomes to unhealthy tissue or to activate events such as, but not limited to, endocytosis. Liposomes may contain a low or a high pH in order to improve the delivery of the pharmaceutical formulations. [0179] The formation of liposomes may depend on the physicochemical characteristics such as, but not limited to, the pharmaceutical formulation entrapped and the liposomal ingredients, the nature of the medium in which the lipid vesicles are dispersed, the effective concentration of the entrapped substance and its potential toxicity, any additional processes involved during the application and/or delivery of the vesicles, the optimization size, polydispersity and the shelf-life of the vesicles for the intended application, and the batch-to- batch reproducibility and possibility of large-scale production of safe and efficient liposomal products. [0180] In some embodiments, pharmaceutical compositions described herein may include, without limitation, liposomes such as those formed from 1,2-dioleyloxy-N,N- dimethylaminopropane (DODMA) liposomes, DiLa2 liposomes from Marina Biotech (Bothell, WA), 1,2-dilinoleyloxy-3-dimethylaminopropane (DLin-DMA), 2,2-dilinoleyl-4-(2- dimethylaminoethyl)-[1,3]-dioxolane (DLin-KC2-DMA), and MC3 (US20100324120; the contents of which are herein incorporated by reference in its entirety) and liposomes which may deliver small molecule drugs such as, but not limited to, DOXIL® from Janssen Biotech, Inc. (Horsham, PA). [0181] In some embodiments, pharmaceutical compositions described herein may include, without limitation, liposomes such as those formed from the synthesis of stabilized plasmid- lipid particles (SPLP) or stabilized nucleic acid lipid particle (SNALP) that have been previously described and shown to be suitable for oligonucleotide delivery in vitro and in vivo (see Wheeler et al. Gene Therapy. 19996:271-281; Zhang et al. Gene Therapy. 1999 6:1438-1447; Jeffs et al. Pharm Res. 200522:362-372; Morrissey et al., Nat Biotechnol. 2005 2:1002-1007; Zimmermann et al., Nature. 2006441:111-114; Heyes et al. J Contr Rel. 2005 107:276-287; Semple et al. Nature Biotech. 201028:172-176; Judge et al. J Clin Invest. 2009 119:661-673; deFougerolles Hum Gene Ther. 200819:125-132; the contents of each of which are incorporated herein in their entireties). The original manufacture method by Wheeler et al. was a detergent dialysis method, which was later improved by Jeffs et al. and is referred to as the spontaneous vesicle formation method. The liposome formulations may be composed of 3 to 4 lipid components in addition to the nucleic acid-based compositions. As a non-limiting example, a liposome can contain, but is not limited to, 55% cholesterol, 20% disteroylphosphatidyl choline (DSPC), 10% PEG-S-DSG, and 15% 1,2-dioleyloxy-N,N- dimethylaminopropane (DODMA), as described by Jeffs et al. In another example, certain liposome formulations may contain, but are not limited to, 48% cholesterol, 20% DSPC, 2% PEG-c-DMA, and 30% cationic lipid, where the cationic lipid can be 1,2-distearloxy-N,N- dimethylaminopropane (DSDMA), DODMA, DLin-DMA, or 1,2-dilinolenyloxy-3- dimethylaminopropane (DLenDMA), as described by Heyes et al. In another example, the nucleic acid-lipid particle may comprise a cationic lipid comprising from about 50 mol % to about 85 mol % of the total lipid present in the particle; a non-cationic lipid comprising from about 13 mol % to about 49.5 mol % of the total lipid present in the particle; and a conjugated lipid that inhibits aggregation of particles comprising from about 0.5 mol % to about 2 mol % of the total lipid present in the particle as described in WO2009127060 to Maclachlan et al, the contents of which are incorporated herein by reference in their entirety. In another example, the nucleic acid-lipid particle may be any nucleic acid-lipid particle disclosed in US2006008910 to Maclachlan et al., the contents of which are incorporated herein by reference in their entirety. As a non-limiting example, the nucleic acid-lipid particle may comprise a cationic lipid of Formula I, a non-cationic lipid, and a conjugated lipid that inhibits aggregation of particles. [0182] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated in a lipid vesicle which may have crosslinks between functionalized lipid bilayers. [0183] In some embodiments, the liposome may contain a sugar-modified lipid disclosed in US5595756 to Bally et al., the contents of which are incorporated herein by reference in their entirety. The lipid may be a ganglioside and cerebroside in an amount of about 10 mol percent. [0184] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated in a liposome comprising a cationic lipid. The liposome may have a molar ratio of nitrogen atoms in the cationic lipid to the phosphates in the nucleic acid-based compositions (N:P ratio) of between 1:1 and 20:1 as described in International Publication No. WO2013006825, the contents of which are herein incorporated by reference in its entirety. In some embodiments, the liposome may have a N:P ratio of greater than 20:1 or less than 1:1. [0185] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated in a lipid-polycation complex. The formation of the lipid- polycation complex may be accomplished by methods known in the art and/or as described in U.S. Pub. No. 20120178702, the contents of which are herein incorporated by reference in its entirety. As a non-limiting example, the polycation may include a cationic peptide or a polypeptide such as, but not limited to, polylysine, polyornithine and/or polyarginine and the cationic peptides described in International Pub. No. WO2012013326; herein incorporated by reference in its entirety. In some embodiments, the nucleic acid-based compositions may be formulated in a lipid-polycation complex which may further include a neutral lipid such as, but not limited to, cholesterol or dioleoyl phosphatidylethanolamine (DOPE). [0186] The liposome formulation may be influenced by, but not limited to, the selection of the cationic lipid component, the degree of cationic lipid saturation, the nature of the PEGylation, ratio of all components and biophysical parameters such as size. In one example by Semple et al. (Semple et al. Nature Biotech. 201028:172-176; the contents of which are herein incorporated by reference in its entirety), the liposome formulation was composed of 57.1 % cationic lipid, 7.1% dipalmitoylphosphatidylcholine, 34.3 % cholesterol, and 1.4% PEG-c-DMA. [0187] In some embodiments, the ratio of PEG in the lipid nanoparticle (LNP) formulations may be increased or decreased and/or the carbon chain length of the PEG lipid may be modified from C14 to C18 to alter the pharmacokinetics and/or biodistribution of the LNP formulations. As a non-limiting example, LNP formulations may contain 1-5% of the lipid molar ratio of PEG-c-DOMG as compared to the cationic lipid, DSPC and cholesterol. In some embodiments, the PEG-c-DOMG may be replaced with a PEG lipid such as, but not limited to, PEG-DSG (1,2-Distearoyl-sn-glycerol, methoxypolyethylene glycol) or PEG-DPG (1,2-Dipalmitoyl-sn-glycerol, methoxypolyethylene glycol). The cationic lipid may be selected from any lipid known in the art such as, but not limited to, DLin-MC3-DMA, DLin- DMA, C12-200 and DLin-KC2-DMA. [0188] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated in a lipid nanoparticle such as the lipid nanoparticles described in International Publication No. WO2012170930, the contents of which are herein incorporated by reference in its entirety. [0189] In some embodiments, the cationic lipid which may be used in formulations of the present disclosure may be selected from, but not limited to, a cationic lipid described in International Publication Nos. WO2012040184, WO2011153120, WO2011149733, WO2011090965, WO2011043913, WO2011022460, WO2012061259, WO2012054365, WO2012044638, WO2010080724, WO201021865 and WO2008103276, US Patent Nos. 7,893,302, 7,404,969 and 8,283,333 and US Patent Publication No. US20100036115 and US20120202871; the contents of each of which is herein incorporated by reference in their entirety. The cationic lipid may be selected from, but not limited to, formula A described in International Publication Nos. WO2012040184, WO2011153120, WO2011149733, WO2011090965, WO2011043913, WO2011022460, WO2012061259, WO2012054365 and WO2012044638; the contents of each of which is herein incorporated by reference in their entirety. Alternatively, the cationic lipid may be selected from, but not limited to, formula CLI-CLXXIX of International Publication No. WO2008103276, formula CLI-CLXXIX of US Patent No. 7,893,302, formula CLI-CLXXXXII of US Patent No. 7,404,969 and formula I-VI of US Patent Publication No. US20100036115; the contents of each of which are herein incorporated by reference in their entirety. The cationic lipid may be a multivalent cationic lipid such as the cationic lipid disclosed in US Patent No. 7223887 to Gaucheron et al., the contents of which are incorporated herein by reference in their entirety. The cationic lipid may have a positively-charged head group including two quaternary amine groups and a hydrophobic portion including four hydrocarbon chains as described in US Patent No. 7223887 to Gaucheron et al., the contents of which are incorporated herein by reference in their entirety. The cationic lipid may be biodegradable as the biodegradable lipids disclosed in US20130195920 to Maier et al., the contents of which are incorporated herein by reference in their entirety. The cationic lipid may have one or more biodegradable groups located in a lipidic moiety of the cationic lipid as described in formula I-IV in US 20130195920 to Maier et al., the contents of which are incorporated herein by reference in their entirety. [0190] As a non-limiting example, the cationic lipid may be selected from (20Z,23Z)- N,N-dimethylnonacosa-20,23-dien-10-amine, (17Z,20Z)-N,N-dimemylhexacosa-17,20-dien- 9-amine, (1Z,19Z)-N5N-dimethylpentacosa-l 6, 19-dien-8-amine, (13Z,16Z)-N,N- dimethyldocosa-13,16-dien-5-amine, (12Z,15Z)-N,N-dimethylhenicosa-12,15-dien-4-amine, (14Z,17Z)-N,N-dimethyltricosa-14,17-dien-6-amine, (15Z,18Z)-N,N-dimethyltetracosa- 15,18-dien-7-amine, (18Z,21Z)-N,N-dimethylheptacosa-18,21-dien-10-amine, (15Ζ,18Ζ)- Ν,Ν-dimethyltetracosa-15,18-dien-5-amine, (14Z,17Z)-N,N-dimethyltricosa-14,17-dien-4- amine, (19Z,22Z)-N,N-dimeihyloctacosa-19,22-dien-9-amine, (18Z,21 Z)-N,N- dimethylheptacosa- 18 ,21 -dien-8 –amine, (17Z,20Z)-N,N-dimethylhexacosa- 17,20-dien-7- amine, (16Z,19Z)-N,N-dimethylpentacosa-16,19-dien-6-amine, (22Z,25Z)-N,N- dimethylhentriaconta-22,25-dien-10-amine, (21 Z ,24Z)-N,N-dimethyltriaconta-21,24-dien-9- amine, (18Z)-N,N-dimetylheptacos-18-en-10-amine, (17Z)-N,N-dimethylhexacos-17-en-9- amine, (19Z,22Z)-N,N-dimethyloctacosa-19,22-dien-7-amine, N,N-dimethylheptacosan-10- amine, (20Z,23Z)-N-ethyl-N-methylnonacosa-20,23-dien-l0-amine, 1-[(11Z,14Z)-l- nonylicosa-11,14-dien-l-yl] pyrrolidine, (20Z)-N,N-dimethylheptacos-20-en-l 0-amine, (15Z)-N,N-dimethyl eptacos-15-en-l 0-amine, (14Z)-N,N-dimethylnonacos-14-en-l0-amine, (17Z)-N,N-dimethylnonacos-17-en-l0-amine, (24Z)-N,N-dimethyltritriacont-24-en-l0-amine, (20Z)-N,N-dimethylnonacos-20-en-l 0-amine, (22Z)-N,N-dimethylhentriacont-22-en-l0- amine, (16Z)-N,N-dimethylpentacos-16-en-8-amine, (12Z,15Z)-N,N-dimethyl-2- nonylhenicosa-12,15-dien-1–amine, (13Z,16Z)-N,N-dimethyl-3-nonyldocosa-l3,16-dien-l– amine, N,N-dimethyl-l-[(lS,2R)-2-octylcyclopropyl] eptadecan-8-amine, 1-[(1S,2R)-2- hexylcyclopropyl]-N,N-dimethylnonadecan-10-amine, Ν,Ν-dimethyl-1-[(1S ,2R)-2- octylcyclopropyl]nonadecan-10-amine, N,N-dimethyl-21-[(lS,2R)-2- octylcyclopropyl]henicosan-l0-amine,Ν,Ν-dimethyl-1-[(1S,2S)-2-{[(lR,2R)-2- pentylcycIopropyl]methyl}cyclopropyl]nonadecan-10-amine,Ν,Ν-dimethyl-1-[(1S,2R)-2- octylcyclopropyl]hexadecan-8-amine, Ν,Ν-dimethyl-[(lR,2S)-2- undecyIcyclopropyl]tetradecan-5-amine, N,N-dimethyl-3-{7-[(1S,2R)-2- octylcyclopropyl]heptyl} dodecan-1–amine, 1-[(1R,2S)-2-hepty lcyclopropyl]-Ν,Ν- dimethyloctadecan-9–amine, 1-[(1S,2R)-2-decylcyclopropyl]-N,N-dimethylpentadecan-6- amine, N,N-dimethyl-l-[(lS,2R)-2-octylcyclopropyl]pentadecan-8-amine, R-N,N-dimethyl-1- [(9Z,12Z)-octadeca-9,12-dien-1-yloxy]-3-(octyloxy)propan-2-amine, S-N,N-dimethyl-1- [(9Z,12Z)-octadeca-9,12-dien-1-yloxy]-3-(octyloxy)propan-2-amine, 1-{2-[(9Z,12Z)- octadeca-9,12-dien-1-yloxy]-1-[(octyloxy)methyl]ethyl}pyrrolidine, (2S)-N,N-dimethyl-1- [(9Z,12Z)-octadeca-9,12-dien-1-yloxy]-3-[(5Z)-oct-5-en-1-yloxy]propan-2-amine, 1-{2- [(9Z,12Z)-octadeca-9,12-dien-1-yloxy]-1-[(octyloxy)methyl]ethyl}azetidine, (2S)-1- (hexyloxy)-N,N-dimethyl-3-[(9Z,12Z)-octadeca-9,12-dien-1-yloxy]propan-2-amine, (2S)-1- (heptyloxy)-N,N-dimethyl-3-[(9Z,12Z)-octadeca-9,12-dien-1-yloxy]propan-2-amine, Ν,Ν- dimethyl-1-(nonyloxy)-3-[(9Z,12Z)-octadeca-9,12-dien-1-yloxy]propan-2-amine, Ν,Ν- dimethyl-1-[(9Z)-octadec-9-en-1-yloxy]-3-(octyloxy)propan-2-amine; (2S)-N,N-dimethyl-1- [(6Z,9Z,12Z)-octadeca-6,9,12-trien-1-yloxy]-3-(octyloxy)propan-2-amine, (2S)-1- [(11Z,14Z)-icosa-11,14-dien-1-yloxy]-N,N-dimethyl-3-(pentyloxy)propan-2-amine, (2S)-1- (hexyloxy)-3-[(11Z,14Z)-icosa-11,14-dien-1-yloxy]-N,N-dimethylpropan-2-amine, 1- [(11Z,14Z)-icosa-11,14-dien-1-yloxy]-Ν,Ν-dimethy1-3-(octyloxy)propan-2-amine, 1- [(13Z,16Z)-docosa-l3,16-dien-l-yloxy]-N,N-dimethyl-3-(octyloxy)propan-2-amine, (2S)-1- [(13Z,16Z)-docosa-13,16-dien-1-yloxy]-3-(hexyloxy)-N,N-dimethylpropan-2-amine, (2S)-1- [(13Z)-docos-13-en-1-yloxy]-3-(hexyloxy)-N,N-dimethylpropan-2-amine, 1-[(13Z)-docos- 13-en-1-yloxy]-N,N-dimethyl-3-(octyloxy)propan-2-amine, 1-[(9Z)-hexadec-9-en-1-yloxy]- N,N-dimethyl-3-(octyloxy)propan-2-amine, (2R)-N,N-dimethyl-H(1-metoylo ctyl)oxy]-3- [(9Z,12Z)-octadeca-9,12-dien-1-yloxy]propan-2-amine, (2R)-1-[(3,7-dimethyloctyl)oxy]- N,N-dimethyl-3-[(9Z,12Z)-octadeca-9,12-dien-1-yloxy]propan-2-amine, N,N-dimethyl-1- (octyloxy)-3-({8-[(1S,2S)-2-{[(1R,2R)-2- pentylcyclopropyl]methyl}cyclopropyl]octyl}oxy)propan-2-amine, N,N-dimethyl-1-{[8-(2- oc1ylcyclopropyl)octyl]oxy}-3-(octyloxy)propan-2-amine and (1lE,20Z,23Z)-N,N- dimethylnonacosa-l1,20,2-trien-10-amine or a pharmaceutically acceptable salt or stereoisomer thereof. [0191] In some embodiments, the lipid may be a cleavable lipid such as those described in International Publication No. WO2012170889, the contents of which are herein incorporated by reference in their entirety. [0192] In some embodiments, the nanoparticles described herein may comprise at least one cationic polymer described herein and/or known in the art. [0193] In some embodiments, the cationic lipid may be synthesized by methods known in the art and/or as described in International Publication Nos. WO2012040184, WO2011153120, WO2011149733, WO2011090965, WO2011043913, WO2011022460, WO2012061259, WO2012054365, WO2012044638, WO2010080724 and WO201021865; the contents of each of which is herein incorporated by reference in their entirety. [0194] In some embodiments, the LNP formulations of the nucleic acid-based compositions may contain PEG-c-DOMG at 3% lipid molar ratio. In some embodiments, the LNP formulations of the nucleic acid-based compositions may contain PEG-c-DOMG at 1.5% lipid molar ratio. [0195] In some embodiments, the pharmaceutical compositions of the nucleic acid-based compositions may include at least one of the PEGylated lipids described in International Publication No. 2012099755, the contents of which is herein incorporated by reference in its entirety. [0196] In some embodiments, the LNP formulation may contain PEG-DMG 2000 (1,2- dimyristoyl-sn-glycero-3-phophoethanolamine-N-[methoxy(polyethylene glycol)-2000). In some embodiments, the LNP formulation may contain PEG-DMG 2000, a cationic lipid known in the art and at least one other component. In some embodiments, the LNP formulation may contain PEG-DMG 2000, a cationic lipid known in the art, DSPC and cholesterol. As a non-limiting example, the LNP formulation may contain PEG-DMG 2000, DLin-DMA, DSPC and cholesterol. As another non-limiting example the LNP formulation may contain PEG-DMG 2000, DLin-DMA, DSPC and cholesterol in a molar ratio of 2:40:10:48 (see e.g., Geall et al., Non-viral delivery of self-amplifying RNA vaccines, PNAS 2012; PMID: 22908294; herein incorporated by reference in its entirety). As another non- limiting example, the nucleic acid-based compositions described herein may be formulated in a nanoparticle to be delivered by a parenteral route as described in U.S. Pub. No. 20120207845; the contents of which is herein incorporated by reference in its entirety. The cationic lipid may also be the cationic lipids disclosed in US20130156845 to Manoharan et al. and US 20130129785 to Manoharan et al., WO 2012047656 to Wasan et al., WO 2010144740 to Chen et al., WO 2013086322 to Ansell et al., or WO 2012016184 to Manoharan et al., the contents of each of which are incorporated herein by reference in their entirety. [0197] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with a plurality of cationic lipids, such as a first and a second cationic lipid as described in US20130017223 to Hope et al., the contents of which are incorporated herein by reference in their entirety. The first cationic lipid can be selected on the basis of a first property and the second cationic lipid can be selected on the basis of a second property, where the properties may be determined as outlined in US20130017223, the contents of which are herein incorporated by reference in its entirety. In some embodiments, the first and second properties are complementary. [0198] The nucleic acid-based compositions described herein may be formulated with a lipid particle comprising one or more cationic lipids and one or more second lipids, and one or more nucleic acids, wherein the lipid particle comprises a solid core, as described in US Patent Publication No. US20120276209 to Cullis et al., the contents of which are incorporated herein by reference in their entirety. [0199] In some embodiments, the nucleic acid-based compositions of the present disclosure may be complexed with a cationic amphiphile in an oil-in-water (o/w) emulsion such as described in EP2298358 to Satishchandran et al., the contents of which are incorporated herein by reference in their entirety. The cationic amphiphile may be a cationic lipid, modified or unmodified spermine, bupivacaine, or benzalkonium chloride and the oil may be a vegetable or an animal oil. As a non-limiting example, at least 10% of the nucleic acid-cationic amphiphile complex is in the oil phase of the oil-in-water emulsion (see e.g., the complex described in European Publication No. EP2298358 to Satishchandran et al., the contents of which are herein incorporated by reference in its entirety). [0200] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with a composition comprising a mixture of cationic compounds and neutral lipids. As a non-limiting example, the cationic compounds may be formula (I) disclosed in WO 1999010390 to Ansell et al., the contents of which are disclosed herein by reference in their entirety, and the neutral lipid may be selected from the group consisting of diacylphosphatidylcholine, diacylphosphatidylethanolamine, ceramide and sphingomyelin. In another non-limiting example, the lipid formulation may comprise a cationic lipid of formula A, a neutral lipid, a sterol and a PEG or PEG-modified lipid disclosed in US 20120101148 to Akinc et al., the contents of which are incorporated herein by reference in their entirety. [0201] In some embodiments, the LNP formulation may be formulated by the methods described in International Publication Nos. WO2011127255 or WO2008103276, each of which are herein incorporated by reference in their entirety. As a non-limiting example, the nucleic acid-based compositions of the present disclosure may be encapsulated in any of the lipid nanoparticle (LNP) formulations described in WO2011127255 and/or WO2008103276; the contents of each of which are herein incorporated by reference in their entirety. [0202] In some embodiments, the LNP formulations described herein may comprise a polycationic composition. As a non-limiting example, the polycationic composition may be selected from formula 1-60 of US Patent Publication No. US20050222064; the contents of which is herein incorporated by reference in its entirety. The LNP formulations comprising a polycationic composition may be used for the delivery of the nucleic acid-based compositions described herein in vivo and/or in vitro. [0203] In some embodiments, the LNP formulations described herein may additionally comprise a permeability enhancer molecule. Non-limiting permeability enhancer molecules are described in US Patent Publication No. US20050222064; the contents of which is herein incorporated by reference in its entirety. [0204] In some embodiments, the pharmaceutical compositions may be formulated in liposomes such as, but not limited to, DiLa2 liposomes (Marina Biotech, Bothell, WA), SMARTICLES®/NOV340 (Marina Biotech, Bothell, WA), neutral DOPC (1,2-dioleoyl-sn- glycero-3-phosphocholine) based liposomes (e.g., siRNA delivery for ovarian cancer (Landen et al. Cancer Biology & Therapy 20065(12)1708-1713); the contents of which is herein incorporated by reference in its entirety) and hyaluronan-coated liposomes (Quiet Therapeutics, Israel). [0205] In some embodiments, the pharmaceutical compositions may be formulated with any amphoteric liposome disclosed in WO 2008/043575 to Panzner and US 8580297 to Essler et al. (Marina Biotech), the contents of which are incorporated herein by reference in their entirety. The amphoteric liposome may comprise a mixture of lipids including a cationic amphiphile, an anionic amphiphile and optional one or more neutral amphiphiles. The amphoteric liposome may comprise amphoteric compounds based on amphiphilic molecules, the head groups of which being substituted with one or more amphoteric groups. In some embodiments, the pharmaceutical compositions may be formulated with an amphoteric lipid comprising one or more amphoteric groups having an isoelectric point between 4 and 9, as disclosed in US 20140227345 to Essler et al. (Marina Biotech), the contents of which are incorporated herein by reference in their entirety. [0206] In some embodiments, the pharmaceutical composition may be formulated with liposomes comprising a sterol derivative as disclosed in US 7312206 to Panzner et al. (Novosom), the contents of which are incorporated herein by reference in their entirety. In some embodiments, the pharmaceutical composition may be formulated with amphoteric liposomes comprising at least one amphipathic cationic lipid, at least one amphipathic anionic lipid, and at least one neutral lipid, or liposomes comprise at least one amphipathic lipid with both a positive and a negative charge, and at least one neutral lipid, wherein the liposomes are stable at pH 4.2 and pH 7.5, as disclosed in US Pat. No. 7780983 to Panzner et al. (Novosom), the contents of which are incorporated herein by reference in their entirety. In some embodiments, the pharmaceutical composition may be formulated with liposomes comprising a serum-stable mixture of lipids taught in US 20110076322 to Panzner et al, the contents of which are incorporated herein by reference in their entirety, capable of encapsulating the nucleic acid-based compositions of the present disclosure. The lipid mixture comprises phosphatidylcholine and phosphatidylethanolamine in a ratio in the range of about 0.5 to about 8. The lipid mixture may also include pH sensitive anionic and cationic amphiphiles, such that the mixture is amphoteric, being negatively charged or neutral at pH 7.4 and positively charged at pH 4. The drug/lipid ratio may be adjusted to target the liposomes to particular organs or other sites in the body. In some embodiments, liposomes loaded with the nucleic acid-based compositions of the present disclosure as cargo, are prepared by the method disclosed in US 20120021042 to Panzner et al., the contents of which are incorporated herein by reference in their entirety. The method comprises steps of admixing an aqueous solution of a polyanionic active agent and an alcoholic solution of one or more amphiphiles and buffering said admixture to an acidic pH, wherein the one or more amphiphiles are susceptible of forming amphoteric liposomes at the acidic pH, thereby to form amphoteric liposomes in suspension encapsulating the active agent. [0207] The nanoparticle formulations may be a carbohydrate nanoparticle comprising a carbohydrate carrier and a nucleic acid-based composition (e.g., an ASO). As a non-limiting example, the carbohydrate carrier may include, but is not limited to, an anhydride-modified phytoglycogen or glycogen-type material, phtoglycogen octenyl succinate, phytoglycogen beta-dextrin, anhydride-modified phytoglycogen beta-dextrin. (See e.g., International Publication No. WO2012109121; the contents of which is herein incorporated by reference in its entirety). [0208] Lipid nanoparticle formulations may be improved by replacing the cationic lipid with a biodegradable cationic lipid which is known as a rapidly eliminated lipid nanoparticle (reLNP). Ionizable cationic lipids, such as, but not limited to, DLinDMA, DLin-KC2-DMA, and DLin-MC3-DMA, have been shown to accumulate in plasma and tissues over time and may be a potential source of toxicity. The rapid metabolism of the rapidly eliminated lipids can improve the tolerability and therapeutic index of the lipid nanoparticles by an order of magnitude from a 1 mg/kg dose to a 10 mg/kg dose in rat. Inclusion of an enzymatically degraded ester linkage can improve the degradation and metabolism profile of the cationic component, while still maintaining the activity of the reLNP formulation. The ester linkage can be internally located within the lipid chain or it may be terminally located at the terminal end of the lipid chain. The internal ester linkage may replace any carbon in the lipid chain. [0209] In some embodiments, the nucleic acid-based compositions may be formulated as a lipoplex, such as, without limitation, the ATUPLEXTM system, the DACC system, the DBTC system and other siRNA-lipoplex technology from Silence Therapeutics (London, United Kingdom), STEMFECTTM from STEMGENT® (Cambridge, MA), and polyethylenimine (PEI) or protamine-based targeted and non-targeted delivery of nucleic acids (Aleku et al. Cancer Res. 200868:9788-9798; Strumberg et al. Int J Clin Pharmacol Ther 201250:76-78; Santel et al., Gene Ther 200613:1222-1234; Santel et al., Gene Ther 200613:1360-1370; Gutbier et al., Pulm Pharmacol. Ther. 201023:334-344; Kaufmann et al. Microvasc Res 2010 80:286-293Weide et al. J Immunother. 200932:498-507; Weide et al. J Immunother. 2008 31:180-188; Pascolo Expert Opin. Biol. Ther. 4:1285-1294; Fotin-Mleczek et al., 2011 J. Immunother. 34:1-15; Song et al., Nature Biotechnol. 2005, 23:709-717; Peer et al., Proc Natl Acad Sci U S A. 20076;104:4095-4100; deFougerolles Hum Gene Ther. 200819:125-132; the contents of each of which are incorporated herein by reference in its entirety). [0210] In some embodiments such formulations may also be constructed or compositions altered such that they passively or actively are directed to different cell types in vivo, including but not limited to hepatocytes, immune cells, tumor cells, endothelial cells, antigen presenting cells, and leukocytes (Akinc et al. Mol Ther. 201018:1357-1364; Song et al., Nat Biotechnol. 200523:709-717; Judge et al., J Clin Invest.2009119:661-673; Kaufmann et al., Microvasc Res 201080:286-293; Santel et al., Gene Ther 200613:1222-1234; Santel et al., Gene Ther 200613:1360-1370; Gutbier et al., Pulm Pharmacol. Ther. 201023:334-344; Basha et al., Mol. Ther. 201119:2186-2200; Fenske and Cullis, Expert Opin Drug Deliv. 20085:25-44; Peer et al., Science. 2008319:627-630; Peer and Lieberman, Gene Ther. 2011 18:1127-1133; the contents of each of which are incorporated herein by reference in its entirety). One example of passive targeting of formulations to liver cells includes the DLin- DMA, DLin-KC2-DMA and DLin-MC3-DMA-based lipid nanoparticle formulations which have been shown to bind to apolipoprotein E and promote binding and uptake of these formulations into hepatocytes in vivo (Akinc et al. Mol Ther. 201018:1357-1364; the contents of which is herein incorporated by reference in its entirety). Formulations can also be selectively targeted through expression of different ligands on their surface as exemplified by, but not limited by, folate, transferrin, N-acetylgalactosamine (GalNAc), and antibody targeted approaches (Kolhatkar et al., Curr Drug Discov Technol. 20118:197-206; Musacchio and Torchilin, Front Biosci. 201116:1388-1412; Yu et al., Mol Membr Biol. 201027:286-298; Patil et al., Crit Rev Ther Drug Carrier Syst. 200825:1-61; Benoit et al., Biomacromolecules. 201112:2708-2714; Zhao et al., Expert Opin Drug Deliv. 20085:309- 319; Akinc et al., Mol Ther. 201018:1357-1364; Srinivasan et al., Methods Mol Biol. 2012 820:105-116; Ben-Arie et al., Methods Mol Biol. 2012757:497-507; Peer 2010 J Control Release. 20:63-68; Peer et al., Proc Natl Acad Sci U S A. 2007104:4095-4100; Kim et al., Methods Mol Biol. 2011721:339-353; Subramanya et al., Mol Ther. 201018:2028-2037; Song et al., Nat Biotechnol. 200523:709-717; Peer et al., Science. 2008319:627-630; Peer and Lieberman, Gene Ther. 201118:1127-1133; the contents of each of which are incorporated herein by reference in its entirety). [0211] In some embodiments, the nucleic acid-based compositions is formulated as a solid lipid nanoparticle. A solid lipid nanoparticle (SLN) may be spherical with an average diameter between 10 to 1000 nm. SLN possess a solid lipid core matrix that can solubilize lipophilic molecules and may be stabilized with surfactants and/or emulsifiers. The lipid nanoparticle may be a self-assembly lipid-polymer nanoparticle (see Zhang et al., ACS Nano, 2008, 2 (8), pp 1696–1702; the contents of which are herein incorporated by reference in its entirety). [0212] In some embodiments, the nucleic acid-based compositions of the present disclosure can be formulated for controlled release and/or targeted delivery. As used herein, “controlled release” refers to a pharmaceutical composition or compound release profile that conforms to a particular pattern of release to affect a therapeutic outcome. In some embodiments, the nucleic acid-based compositions may be encapsulated into a delivery agent described herein and/or known in the art for controlled release and/or targeted delivery. As used herein, the term “encapsulate” means to enclose, surround or encase. As it relates to the formulation of the compounds of the disclosure, encapsulation may be substantial, complete or partial. The term “substantially encapsulated” means that at least greater than 50, 60, 70, 80, 85, 90, 95, 96, 97, 98, 99, 99.9, 99.9 or greater than 99.999% of the pharmaceutical composition or compound of the disclosure may be enclosed, surrounded or encased within the delivery agent. “Partially encapsulated” means that less than 10, 10, 20, 30, 4050 or less of the pharmaceutical composition or compound of the disclosure may be enclosed, surrounded or encased within the delivery agent. Advantageously, encapsulation may be determined by measuring the escape or the activity of the pharmaceutical composition or compound of the disclosure using fluorescence and/or electron micrograph. For example, at least 1, 5, 10, 20, 30, 40, 50, 60, 70, 80, 85, 90, 95, 96, 97, 98, 99, 99.9, 99.99 or greater than 99.99% of the pharmaceutical composition or compound of the disclosure are encapsulated in the delivery agent. [0213] The nucleic acid-based compositions may be encapsulated into a lipid nanoparticle or a rapidly eliminated lipid nanoparticle and the lipid nanoparticles or a rapidly eliminated lipid nanoparticle may then be encapsulated into a polymer, hydrogel and/or surgical sealant described herein and/or known in the art. As a non-limiting example, the polymer, hydrogel or surgical sealant may be PLGA, ethylene vinyl acetate (EVAc), poloxamer, GELSITE® (Nanotherapeutics, Inc. Alachua, FL), HYLENEX® (Halozyme Therapeutics, San Diego CA), surgical sealants such as fibrinogen polymers (Ethicon Inc. Cornelia, GA), TISSELL® (Baxter International, Inc., Deerfield, IL), PEG-based sealants, and COSEAL® (Baxter International, Inc., Deerfield, IL). [0214] In some embodiments, the lipid nanoparticle may be encapsulated into any polymer known in the art which may form a gel when injected into a subject. As another non- limiting example, the lipid nanoparticle may be encapsulated into a polymer matrix which may be biodegradable. [0215] In some embodiments, the nucleic acid-based compositions formulation for controlled release and/or targeted delivery may also include at least one controlled release coating. Controlled release coatings include, but are not limited to, OPADRY®, polyvinylpyrrolidone/vinyl acetate copolymer, polyvinylpyrrolidone, hydroxypropyl methylcellulose, hydroxypropyl cellulose, hydroxyethyl cellulose, EUDRAGIT RL®, EUDRAGIT RS® and cellulose derivatives such as ethylcellulose aqueous dispersions (AQUACOAT® and SURELEASE®). [0216] In some embodiments, the controlled release and/or targeted delivery formulation may comprise at least one degradable polyester which may contain polycationic side chains. Degradable polyesters include, but are not limited to, poly(serine ester), poly(L-lactide-co-L- lysine), poly(4-hydroxy-L-proline ester), and combinations thereof. In some embodiments, the degradable polyesters may include a PEG conjugation to form a PEGylated polymer. [0217] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with a targeting lipid with a targeting moiety such as the targeting moieties disclosed in US20130202652 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety. As a non-limiting example, the targeting moiety of formula I of US 20130202652 to Manoharan et al. may selected in order to favor the lipid being localized with a desired organ, tissue, cell, cell type or subtype, or organelle. Non-limiting targeting moieties that are contemplated in the present disclosure include transferrin, anisamide, an RGD peptide, prostate specific membrane antigen (PSMA), fucose, an antibody, or an aptamer. [0218] In some embodiments, the nucleic acid-based compositions of the present disclosure may be encapsulated in a therapeutic nanoparticle. Therapeutic nanoparticles may be formulated by methods described herein and known in the art such as, but not limited to, International Pub Nos. WO2010005740, WO2010030763, WO2010005721, WO2010005723, WO2012054923, US Pub. Nos. US20110262491, US20100104645, US20100087337, US20100068285, US20110274759, US20100068286 and US20120288541 and US Pat No. 8,206,747, 8,293,276, 8,318,208 and 8,318,211; the contents of each of which are herein incorporated by reference in their entirety. Therapeutic polymer nanoparticles may be identified by the methods described in US Pub No. US20120140790, the contents of which are herein incorporated by reference in its entirety. [0219] In some embodiments, the therapeutic nanoparticle may be formulated for sustained release. As used herein, “sustained release” refers to a pharmaceutical composition or compound that conforms to a release rate over a specific period of time. The period of time may include, but is not limited to, hours, days, weeks, months and years. As a non-limiting example, the sustained release nanoparticle may comprise a polymer and a therapeutic agent such as, but not limited to, the nucleic acid-based compositions of the present disclosure (see International Pub No. 2010075072 and US Pub No. US20100216804, US20110217377 and US20120201859, the contents of each of which are herein incorporated by reference in their entirety). [0220] In some embodiments, the therapeutic nanoparticles may be formulated to be target specific. As a non-limiting example, the therapeutic nanoparticles may include a corticosteroid (see International Pub. No. WO2011084518; the contents of which are herein incorporated by reference in its entirety). In some embodiments, the therapeutic nanoparticles may be formulated to be cancer specific. As a non-limiting example, the therapeutic nanoparticles may be formulated in nanoparticles described in International Pub No. WO2008121949, WO2010005726, WO2010005725, WO2011084521 and US Pub No. US20100069426, US20120004293 and US20100104655, the contents of each of which are herein incorporated by reference in their entirety. [0221] In some embodiments, the nanoparticles of the present disclosure may comprise a polymeric matrix. As a non-limiting example, the nanoparticle may comprise two or more polymers such as, but not limited to, polyethylenes, polycarbonates, polyanhydrides, polyhydroxyacids, polypropylfumerates, polycaprolactones, polyamides, polyacetals, polyethers, polyesters, poly(orthoesters), polycyanoacrylates, polyvinyl alcohols, polyurethanes, polyphosphazenes, polyacrylates, polymethacrylates, polycyanoacrylates, polyureas, polystyrenes, polyamines, polylysine, poly(ethylene imine), poly(serine ester), poly(L-lactide-co-L-lysine), poly(4-hydroxy-L-proline ester) or combinations thereof. [0222] In some embodiments, the therapeutic nanoparticle comprises a diblock copolymer. In some embodiments, the diblock copolymer may include PEG in combination with a polymer such as, but not limited to, polyethylenes, polycarbonates, polyanhydrides, polyhydroxyacids, polypropylfumerates, polycaprolactones, polyamides, polyacetals, polyethers, polyesters, poly(orthoesters), polycyanoacrylates, polyvinyl alcohols, polyurethanes, polyphosphazenes, polyacrylates, polymethacrylates, polycyanoacrylates, polyureas, polystyrenes, polyamines, polylysine, poly(ethylene imine), poly(serine ester), poly(L-lactide-co-L-lysine), poly(4-hydroxy-L-proline ester) or combinations thereof. [0223] As a non-limiting example, the therapeutic nanoparticle comprises a PLGA-PEG block copolymer (see US Pub. No. US20120004293 and US Pat No. 8,236,330, each of which is herein incorporated by reference in their entirety). In another non-limiting example, the therapeutic nanoparticle is a stealth nanoparticle comprising a diblock copolymer of PEG and PLA or PEG and PLGA (see US Pat No 8,246,968 and International Publication No. WO2012166923, the contents of each of which is herein incorporated by reference in its entirety). [0224] In some embodiments, the therapeutic nanoparticle may comprise a multiblock copolymer such as, but not limited to the multiblock copolymers described in U.S. Pat. No. 8,263,665 and 8,287,910; the contents of each of which are herein incorporated by reference in its entirety. [0225] In some embodiments, the block copolymers described herein may be included in a polyion complex comprising a non-polymeric micelle and the block copolymer. (See e.g., U.S. Pub. No. 20120076836; the contents of which are herein incorporated by reference in its entirety). [0226] In some embodiments, the therapeutic nanoparticle may comprise at least one acrylic polymer. Acrylic polymers include but are not limited to, acrylic acid, methacrylic acid, acrylic acid and methacrylic acid copolymers, methyl methacrylate copolymers, ethoxyethyl methacrylates, cyanoethyl methacrylate, amino alkyl methacrylate copolymer, poly(acrylic acid), poly(methacrylic acid), polycyanoacrylates and combinations thereof. [0227] In some embodiments, the therapeutic nanoparticles may comprise at least one amine-containing polymer such as, but not limited to polylysine, polyethylene imine, poly(amidoamine) dendrimers, poly(beta-amino esters) (See e.g., U.S. Pat. No. 8,287,849; the contents of which are herein incorporated by reference in its entirety) and combinations thereof. [0228] In some embodiments, the therapeutic nanoparticles may comprise at least one degradable polyester which may contain polycationic side chains. Degradable polyesters include, but are not limited to, poly(serine ester), poly(L-lactide-co-L-lysine), poly(4- hydroxy-L-proline ester), and combinations thereof. The degradable polyesters may include a PEG conjugation to form a PEGylated polymer. [0229] In some embodiments, the therapeutic nanoparticle may include a conjugation of at least one targeting ligand. The targeting ligand may be any ligand known in the art such as, but not limited to, a monoclonal antibody. (Kirpotin et al, Cancer Res. 200666:6732-6740; the contents of which are herein incorporated by reference in its entirety). [0230] In some embodiments, the therapeutic nanoparticle may be formulated in an aqueous solution which may be used to target cancer (see International Pub No. WO2011084513 and US Pub No. US20110294717, the contents of each of which is herein incorporated by reference in their entirety). [0231] In some embodiments, the nucleic acid-based compositions may be encapsulated in, linked to and/or associated with synthetic nanocarriers. Synthetic nanocarriers include, but are not limited to, those described in International Pub. Nos. WO2010005740, WO2010030763, WO201213501, WO2012149252, WO2012149255, WO2012149259, WO2012149265, WO2012149268, WO2012149282, WO2012149301, WO2012149393, WO2012149405, WO2012149411, WO2012149454 and WO2013019669, and US Pub. Nos. US20110262491, US20100104645, US20100087337 and US20120244222, the contents of each of which are herein incorporated by reference in their entirety. The synthetic nanocarriers may be formulated using methods known in the art and/or described herein. As a non-limiting example, the synthetic nanocarriers may be formulated by the methods described in International Pub Nos. WO2010005740, WO2010030763 and WO201213501and US Pub. Nos. US20110262491, US20100104645, US20100087337 and US2012024422, the contents of each of which are herein incorporated by reference in their entirety. The synthetic nanocarrier formulations may be lyophilized by methods described in International Pub. No. WO2011072218 and US Pat No. 8,211,473; the contents of each of which are herein incorporated by reference in their entirety. [0232] In some embodiments, the synthetic nanocarriers may contain reactive groups to release the nucleic acid-based compositions described herein (see International Pub. No. WO20120952552 and US Pub No. US20120171229, the contents of each of which are herein incorporated by reference in their entirety). [0233] In some embodiments, the synthetic nanocarriers may be formulated for targeted release. In some embodiments, the synthetic nanocarrier may be formulated to release the nucleic acid-based compositions at a specified pH and/or after a desired time interval. As a non-limiting example, the synthetic nanoparticle may be formulated to release the nucleic acid-based compositions after 24 hours and/or at a pH of 4.5 (see International Pub. Nos. WO2010138193 and WO2010138194 and US Pub Nos. US20110020388 and US20110027217, the contents of each of which is herein incorporated by reference in their entireties). [0234] In some embodiments, the synthetic nanocarriers may be formulated for controlled and/or sustained release of the nucleic acid-based compositions described herein. As a non- limiting example, the synthetic nanocarriers for sustained release may be formulated by methods known in the art, described herein and/or as described in International Pub No. WO2010138192 and US Pub No. 20100303850, the contents each of which is herein incorporated by reference in their entirety. [0235] In some embodiments, the nanoparticle may be optimized for oral administration. The nanoparticle may comprise at least one cationic biopolymer such as, but not limited to, chitosan or a derivative thereof. As a non-limiting example, the nanoparticle may be formulated by the methods described in U.S. Pub. No. 20120282343; the contents of which are herein incorporated by reference in its entirety. [0236] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated in a modular composition such as described in US 8575123 to Manoharan et al., the contents of which are herein incorporated by reference in their entirety. As a non-limiting example, the modular composition may comprise a nucleic acid, e.g., the nucleic acid-based compositions of the present disclosure, at least one endosomolytic component, and at least one targeting ligand. The modular composition may have a formula such as any formula described in US 8575123 to Manoharan et al., the contents of which are herein incorporated by reference in their entirety. [0237] In some embodiments, the nucleic acid-based compositions of the present disclosure may be encapsulated in the lipid formulation to form a stable nucleic acid-lipid particle (SNALP) such as described in US8546554 to de Fougerolles et al., the contents of which are incorporated here by reference in their entirety. The lipid may be cationic or non- cationic. In one non-limiting example, the lipid to nucleic acid ratio (mass/mass ratio) (e.g., lipid to nucleic acid-based compositions ratio) will be in the range of from about 1:1 to about 50:1, from about 1:1 to about 25:1, from about 3:1 to about 15:1, from about 4:1 to about 10:1, from about 5:1 to about 9:1, or about 6:1 to about 9:1, or 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, or 11:1. In another example, the SNALP includes 40% 2,2-Dilinoleyl-4-dimethylaminoethyl- [1,3]-dioxolane (Lipid A), 10% dioleoylphosphatidylcholine (DSPC), 40% cholesterol, 10% polyethylene glycol (PEG)-C-DOMG (mole percent) with a particle size of 63.0±20 nm and a 0.027 nucleic acid/lipid ratio. The nucleic acid-based compositions of the present disclosure may be formulated with a nucleic acid-lipid particle comprising an endosomal membrane destabilizer as disclosed in US 7189705 to Lam et al., the contents of which are incorporated herein by reference in their entirety. As a non-limiting example, the endosomal membrane destabilizer may be a Ca2+ ion. [0238] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with formulated lipid particles (FLiPs) disclosed in US 8148344 to Akinc et al., the contents of which are herein incorporated by reference in their entirety. Akinc et al. teach that FLiPs may comprise at least one of a single or double- stranded oligonucleotide, where the oligonucleotide has been conjugated to a lipophile and at least one of an emulsion or liposome to which the conjugated oligonucleotide has been aggregated, admixed or associated. These particles have surprisingly been shown to effectively deliver oligonucleotides to heart, lung and muscle disclosed in US 8148344 to Akinc et al., the contents of which are herein incorporated by reference in their entirety. [0239] In some embodiments, the nucleic acid-based compositions of the present disclosure may be delivered to a cell using a composition comprising an expression vector in a lipid formulation as described in US 6086913 to Tam et al., the contents of which are incorporated herein by reference in their entirety. The composition disclosed by Tam is serum-stable and comprises an expression vector comprising first and second inverted repeated sequences from an adeno associated virus (AAV), a rep gene from AAV, and a nucleic acid fragment. The expression vector in Tam is complexed with lipids. [0240] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with a lipid formulation disclosed in US 20120270921 to de Fougerolles et al., the contents of which are incorporated herein by reference in their entirety. In one non-limiting example, the lipid formulation may include a cationic lipid having the formula A described in US 20120270921, the contents of which are herein incorporated by reference in its entirety. In another non-limiting example, the compositions of exemplary nucleic acid-lipid particles disclosed in Table A of US20120270921, the contents of which are incorporated herein by reference in their entirety, may be used with the nucleic acid-based compositions of the present disclosure. [0241] In some embodiments, the nucleic acid-based compositions of the present disclosure may be fully encapsulated in a lipid particle disclosed in US 20120276207 to Maurer et al., the contents of which are incorporated herein by reference in their entirety. The particles may comprise a lipid composition comprising preformed lipid vesicles, a charged therapeutic agent, and a destabilizing agent to form a mixture of preformed vesicles and therapeutic agent in a destabilizing solvent, wherein the destabilizing solvent is effective to destabilize the membrane of the preformed lipid vesicles without disrupting the vesicles. [0242] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with a conjugated lipid. In a non-limiting example, the conjugated lipid may have a formula such as described in US 20120264810 to Lin et al., the contents of which are incorporated herein by reference in their entirety. The conjugate lipid may form a lipid particle which further comprises a cationic lipid, a neutral lipid, and a lipid capable of reducing aggregation. [0243] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated in a neutral liposomal formulation such as disclosed in US 20120244207 to Fitzgerald et al., the contents of which are incorporated herein by reference in their entirety. The phrase “neutral liposomal formulation” refers to a liposomal formulation with a near neutral or neutral surface charge at a physiological pH. Physiological pH can be, e.g., about 7.0 to about 7.5, or, e.g., about 7.5, or, e.g., 7.0, 7.1, 7.2, 7.3, 7.4, or 7.5, or, e.g., 7.3, or, e.g., 7.4. An example of a neutral liposomal formulation is an ionizable lipid nanoparticle (iLNP). A neutral liposomal formulation can include an ionizable cationic lipid, e.g., DLin-KC2-DMA. [0244] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with a charged lipid or an amino lipid. As used herein, the term "charged lipid" is meant to include those lipids having one or two fatty acyl or fatty alkyl chains and a quaternary amino head group. The quaternary amine carries a permanent positive charge. The head group can optionally include an ionizable group, such as a primary, secondary, or tertiary amine that may be protonated at physiological pH. The presence of the quaternary amine can alter the pKa of the ionizable group relative to the pKa of the group in a structurally similar compound that lacks the quaternary amine (e.g., the quaternary amine is replaced by a tertiary amine) In some embodiments, a charged lipid is referred to as an "amino lipid." In a non-limiting example, the amino lipid may be any amino lipid described in US20110256175 to Hope et al., the contents of which are incorporated herein by reference in their entirety. For example, the amino lipids may have the structure disclosed in Tables 3-7 of Hope, such as structure (II), DLin-K-C2-DMA, DLin-K2-DMA, DLin-K6- DMA, etc. The resulting pharmaceutical preparations may be lyophilized according to Hope. In another non-limiting example, the amino lipids may be any amino lipid described in US 20110117125 to Hope et al., the contents of which are incorporated herein by reference in their entirety, such as a lipid of structure (I), DLin-K-DMA, DLin-C-DAP, DLin-DAC, DLin-MA, DLin-S-DMA, etc. In another non-limiting example, the amino lipid may have the structure (I), (II), (III), or (IV), or 4-(R)-DUn-K-DMA (VI), 4-(S)-DUn-K-DMA (V) as described in WO2009132131 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety. In another non-limiting example, the charged lipid used in any of the formulations described herein may be any charged lipid described in EP2509636 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety. [0245] In some embodiments, the nucleic acid-based composition s of the present disclosure may be formulated with an association complex containing lipids, liposomes, or lipoplexes. In a non-limiting example, the association complex comprises one or more compounds each having a structure defined by formula (I), a PEG-lipid having a structure defined by formula (XV), a steroid and a nucleic acid disclosed in US8034376 to Manoharan et al., the contents of which are incorporated herein by reference in their entirety. The nucleic acid-based compositions may be formulated with any association complex described in US8034376, the contents of which are herein incorporated by reference in its entirety. [0246] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with reverse head group lipids. As a non-limiting example, the nucleic acid-based compositions may be formulated with a zwitterionic lipid comprising a headgroup wherein the positive charge is located near the acyl chain region and the negative charge is located at the distal end of the head group, such as a lipid having structure (A) or structure (I) described in WO2011056682 to Leung et al., the contents of which are incorporated herein by reference in their entirety. [0247] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated in a lipid bilayer carrier. As a non-limiting example, the nucleic acid-based compositions may be combined with a lipid-detergent mixture comprising a lipid mixture of an aggregation-preventing agent in an amount of about 5 mol% to about 20 mol%, a cationic lipid in an amount of about 0.5 mol% to about 50 mol%, and a fusogenic lipid and a detergent, to provide a nucleic acid-lipid-detergent mixture; and then dialyzing the nucleic acid-lipid-detergent mixture against a buffered salt solution to remove the detergent and to encapsulate the nucleic acid in a lipid bilayer carrier and provide a lipid bilayer-nucleic acid composition, wherein the buffered salt solution has an ionic strength sufficient to encapsulate of from about 40 % to about 80 % of the nucleic acid, described in WO1999018933 to Cullis et al., the contents of which are incorporated herein by reference in their entirety. [0248] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated in a nucleic acid-lipid particle capable of selectively targeting the nucleic acid-based compositions to a kidney, heart, liver, or tumor tissue site. For example, the nucleic acid-lipid particle may comprise (a) a nucleic acid; (b) 1.0 mole % to 45 mole % of a cationic lipid; (c) 0,0 mole % to 90 mole % of another lipid; (d) 1,0 mole % to 10 mole % of a bilayer stabilizing component; (e) 0,0 mole % to 60 mole % cholesterol; and (f) 0,0 mole % to 10 mole % of cationic polymer lipid as described in EP1328254 to Cullis et al., the contents of which are incorporated herein by reference in their entirety. Cullis teaches that varying the amount of each of the cationic lipid, bilayer stabilizing component, another lipid, cholesterol, and cationic polymer lipid can impart tissue selectivity for heart, liver, or tumor tissue site, thereby identifying a nucleic acid-lipid particle capable of selectively targeting a nucleic acid to the heart, liver, or tumor tissue site. Polymers, Biodegradable Nanoparticles, and Core-Shell Nanoparticles [0249] The nucleic acid-based compositions of the disclosure can be formulated using natural and/or synthetic polymers. Non-limiting examples of polymers which may be used for delivery include, but are not limited to, DYNAMIC POLYCONJUGATE® (Arrowhead Research Corp., Pasadena, CA) formulations from MIRUS® Bio (Madison, WI) and Roche Madison (Madison, WI), PHASERXTM polymer formulations such as, without limitation, SMARTT POLYMER TECHNOLOGY™ (PHASERX®, Seattle, WA), DMRI/DOPE, poloxamer, VAXFECTIN® adjuvant from Vical (San Diego, CA), chitosan, cyclodextrin from Calando Pharmaceuticals (Pasadena, CA), dendrimers and poly(lactic-co-glycolic acid) (PLGA) polymers. RONDELTM (RNAi/Oligonucleotide Nanoparticle Delivery) polymers (Arrowhead Research Corporation, Pasadena, CA) and pH responsive co-block polymers such as, but not limited to, PHASERX® (Seattle, WA). [0250] A non-limiting example of chitosan formulation includes a core of positively charged chitosan and an outer portion of negatively charged substrate (U.S. Pub. No. 20120258176; herein incorporated by reference in its entirety). Chitosan includes, but is not limited to N-trimethyl chitosan, mono-N-carboxymethyl chitosan (MCC), N-palmitoyl chitosan (NPCS), EDTA-chitosan, low molecular weight chitosan, chitosan derivatives, or combinations thereof. [0251] In some embodiments, the polymers used in the present disclosure have undergone processing to reduce and/or inhibit the attachment of unwanted substances such as, but not limited to, bacteria, to the surface of the polymer. The polymer may be processed by methods known and/or described in the art and/or described in International Pub. No. WO2012150467, herein incorporated by reference in its entirety. [0252] A non-limiting example of PLGA formulations include, but are not limited to, PLGA injectable depots (e.g., ELIGARD® which is formed by dissolving PLGA in 66% N- methyl-2-pyrrolidone (NMP) and the remainder being aqueous solvent and leuprolide. Once injected, the PLGA and leuprolide peptide precipitates into the subcutaneous space). [0253] Many of these polymer approaches have demonstrated efficacy in delivering oligonucleotides in vivo into the cell cytoplasm (reviewed in de Fougerolles Hum Gene Ther. 200819:125-132; herein incorporated by reference in its entirety). Two polymer approaches that have yielded robust in vivo delivery of nucleic acids, i.e., in the case of small interfering RNA (siRNA), are dynamic polyconjugates and cyclodextrin-based nanoparticles. The first of these delivery approaches uses dynamic polyconjugates and has been shown in vivo in mice to effectively deliver siRNA and silence endogenous target mRNA in hepatocytes (Rozema et al., Proc Natl Acad Sci U S A. 2007104:12982-12887; herein incorporated by reference in its entirety). This particular approach is a multicomponent polymer system whose key features include a membrane-active polymer to which nucleic acid, in this case siRNA, is covalently coupled via a disulfide bond and where both PEG (for charge masking) and N-acetylgalactosamine (for hepatocyte targeting) groups are linked via pH-sensitive bonds (Rozema et al., Proc Natl Acad Sci U S A. 2007104:12982-12887; herein incorporated by reference in its entirety). On binding to the hepatocyte and entry into the endosome, the polymer complex disassembles in the low-pH environment, with the polymer exposing its positive charge, leading to endosomal escape and cytoplasmic release of the siRNA from the polymer. Through replacement of the N-acetylgalactosamine group with a mannose group, it was shown one could alter targeting from asialoglycoprotein receptor-expressing hepatocytes to sinusoidal endothelium and Kupffer cells. Another polymer approach involves using transferrin-targeted cyclodextrin-containing polycation nanoparticles. These nanoparticles have demonstrated targeted silencing of the EWS-FLI1 gene product in transferrin receptor- expressing Ewing’s sarcoma tumor cells (Hu-Lieskovan et al., Cancer Res.200565: 8984- 8982; herein incorporated by reference in its entirety) and siRNA formulated in these nanoparticles was well tolerated in non-human primates (Heidel et al., Proc Natl Acad Sci USA 2007104:5715-21; herein incorporated by reference in its entirety). Both of these delivery strategies incorporate rational approaches using both targeted delivery and endosomal escape mechanisms. [0254] The polymer formulation can permit the sustained or delayed release of nucleic acid-based compositions (e.g., following intramuscular, subcutaneous, intraparenchymal, intrathecal, intracerebroventricular administration). The altered release profile for the nucleic acid-based compositions can result in, for example, translation of an encoded protein over an extended period of time. Biodegradable polymers have been previously used to protect nucleic acids from degradation and been shown to result in sustained release of payloads in vivo (Rozema et al., Proc Natl Acad Sci U S A. 2007104:12982-12887; Sullivan et al., Expert Opin Drug Deliv. 20107:1433-1446; Convertine et al., Biomacromolecules. 2010 Oct 1; Chu et al., Acc Chem Res. 2012 Jan 13; Manganiello et al., Biomaterials. 201233:2301- 2309; Benoit et al., Biomacromolecules. 201112:2708-2714; Singha et al., Nucleic Acid Ther. 20112:133-147; de Fougerolles Hum Gene Ther. 200819:125-132; Schaffert and Wagner, Gene Ther. 200816:1131-1138; Chaturvedi et al., Expert Opin Drug Deliv. 2011 8:1455-1468; Davis, Mol Pharm. 20096:659-668; Davis, Nature 2010464:1067-1070; each of which is herein incorporated by reference in its entirety). [0255] In some embodiments, the pharmaceutical compositions may be sustained release formulations. In further embodiments, the sustained release formulations may be for subcutaneous delivery. Sustained release formulations may include, but are not limited to, PLGA microspheres, ethylene vinyl acetate (EVAc), poloxamer, GELSITE® (Nanotherapeutics, Inc. Alachua, FL), HYLENEX® (Halozyme Therapeutics, San Diego CA), surgical sealants such as fibrinogen polymers (Ethicon Inc. Cornelia, GA), TISSELL® (Baxter International, Inc Deerfield, IL), PEG-based sealants, and COSEAL® (Baxter International, Inc Deerfield, IL). [0256] As a non-limiting example nucleic acid-based compositions may be formulated in PLGA microspheres by preparing the PLGA microspheres with tunable release rates (e.g., days and weeks) and encapsulating the nucleic acid-based compositions in the PLGA microspheres while maintaining the integrity of the nucleic acid-based compositions during the encapsulation process. EVAc are non-biodegradable, biocompatible polymers which are used extensively in pre-clinical sustained release implant applications (e.g., extended release products Ocusert a pilocarpine ophthalmic insert for glaucoma or progestasert a sustained release progesterone intrauterine device; transdermal delivery systems Testoderm, Duragesic and Selegiline; catheters). Poloxamer F-407 NF is a hydrophilic, non-ionic surfactant triblock copolymer of polyoxyethylene-polyoxypropylene-polyoxyethylene having a low viscosity at temperatures less than 5ºC and forms a solid gel at temperatures greater than 15ºC. PEG- based surgical sealants comprise two synthetic PEG components mixed in a delivery device which can be prepared in one minute, seals in 3 minutes and is reabsorbed within 30 days. GELSITE® and natural polymers are capable of in-situ gelation at the site of administration. They have been shown to interact with protein and peptide therapeutic candidates through ionic interaction to provide a stabilizing effect. [0257] Polymer formulations can also be selectively targeted through expression of different ligands as exemplified by, but not limited by, folate, transferrin, and N- acetylgalactosamine (GalNAc) (Benoit et al., Biomacromolecules. 201112:2708-2714; Rozema et al., Proc Natl Acad Sci U S A. 2007104:12982-12887; Davis, Mol Pharm. 2009 6:659-668; Davis, Nature 2010464:1067-1070; each of which is herein incorporated by reference in its entirety). [0258] The nucleic acid-based compositions of the disclosure may be formulated with or in a polymeric compound. The polymer may include at least one polymer such as, but not limited to, polyethenes, polyethylene glycol (PEG), poly(l-lysine)(PLL), PEG grafted to PLL, cationic lipopolymer, biodegradable cationic lipopolymer, polyethyleneimine (PEI), cross- linked branched poly(alkylene imines), a polyamine derivative, a modified poloxamer, a biodegradable polymer, elastic biodegradable polymer, biodegradable block copolymer, biodegradable random copolymer, biodegradable polyester copolymer, biodegradable polyester block copolymer, biodegradable polyester block random copolymer, multiblock copolymers, linear biodegradable copolymer, poly[α-(4-aminobutyl)-L-glycolic acid) (PAGA), biodegradable cross-linked cationic multi-block copolymers, polycarbonates, polyanhydrides, polyhydroxyacids, polypropylfumerates, polycaprolactones, polyamides, polyacetals, polyethers, polyesters, poly(orthoesters), polycyanoacrylates, polyvinyl alcohols, polyurethanes, polyphosphazenes, polyacrylates, polymethacrylates, polycyanoacrylates, polyureas, polystyrenes, polyamines, polylysine, poly(ethylene imine), poly(serine ester), poly(L-lactide-co-L-lysine), poly(4-hydroxy-L-proline ester), acrylic polymers, amine- containing polymers, dextran polymers, dextran polymer derivatives or combinations thereof. [0259] As a non-limiting example, the nucleic acid-based compositions of the disclosure may be formulated with the polymeric compound of PEG grafted with PLL as described in U.S. Pat. No. 6,177,274; herein incorporated by reference in its entirety. The formulation may be used for transfecting cells in vitro or for in vivo delivery of the nucleic acid-based compositions. In another example, the nucleic acid-based compositions may be suspended in a solution or medium with a cationic polymer, in a dry pharmaceutical composition or in a solution that is capable of being dried as described in U.S. Pub. Nos. 20090042829 and 20090042825; each of which are herein incorporated by reference in their entireties. [0260] As another non-limiting example the nucleic acid-based compositions of the disclosure may be formulated with a PLGA-PEG block copolymer (see US Pub. No. US20120004293 and US Pat No. 8,236,330, herein incorporated by reference in their entireties) or PLGA-PEG-PLGA block copolymers (See U.S. Pat. No. 6,004,573, herein incorporated by reference in its entirety). As a non-limiting example, the nucleic acid-based compositions of the disclosure may be formulated with a diblock copolymer of PEG and PLA or PEG and PLGA (see US Pat No 8,246,968, herein incorporated by reference in its entirety). [0261] A polyamine derivative may be used to deliver nucleic acids or to treat and/or prevent a disease or to be included in an implantable or injectable device (U.S. Pub. No. 20100260817 herein incorporated by reference in its entirety). As a non-limiting example, a pharmaceutical composition may include the nucleic acid-based compositions and the polyamine derivative described in U.S. Pub. No. 20100260817, the contents of which are incorporated herein by reference in their entirety. As a non-limiting example the nucleic acid- based compositions of the present disclosure may be delivered using a polyamide polymer such as, but not limited to, a polymer comprising a 1,3-dipolar addition polymer prepared by combining a carbohydrate diazide monomer with a dialkyne unite comprising oligoamines (U.S. Pat. No. 8,236,280; herein incorporated by reference in its entirety). [0262] In some embodiments, the nucleic acid-based compositions of the present disclosure may be formulated with at least one polymer and/or derivatives thereof described in International Publication Nos. WO2011115862, WO2012082574 and WO2012068187 and U.S. Pub. No. 20120283427, the contents of each of which are herein incorporated by reference in their entireties. The nucleic acid-based compositions of the present disclosure may be formulated with a polymer of formula Z as described in WO2011115862, herein incorporated by reference in its entirety. The nucleic acid-based compositions may be formulated with a polymer of formula Z, Z’ or Z’’ as described in International Pub. Nos. WO2012082574 or WO2012068187 and U.S. Pub. No. 2012028342, the contents of each of which are herein incorporated by reference in their entireties. The polymers formulated with the nucleic acid-based compositions of the present disclosure may be synthesized by the methods described in International Pub. Nos. WO2012082574 or WO2012068187, the contents of each of which are herein incorporated by reference in their entireties. [0263] The nucleic acid-based compositions of the disclosure may be formulated with at least one acrylic polymer. Acrylic polymers include but are not limited to, acrylic acid, methacrylic acid, acrylic acid and methacrylic acid copolymers, methyl methacrylate copolymers, ethoxyethyl methacrylates, cyanoethyl methacrylate, amino alkyl methacrylate copolymer, poly(acrylic acid), poly(methacrylic acid), polycyanoacrylates and combinations thereof. [0264] Formulations of nucleic acid-based compositions of the disclosure may include at least one amine-containing polymer such as, but not limited to polylysine, polyethylene imine, poly(amidoamine) dendrimers or combinations thereof. [0265] For example, the nucleic acid-based compositions of the disclosure may be formulated in a pharmaceutical compound including a poly(alkylene imine), a biodegradable cationic lipopolymer, a biodegradable block copolymer, a biodegradable polymer, or a biodegradable random copolymer, a biodegradable polyester block copolymer, a biodegradable polyester polymer, a biodegradable polyester random copolymer, a linear biodegradable copolymer, PAGA, a biodegradable cross-linked cationic multi-block copolymer or combinations thereof. The biodegradable cationic lipopolymer may be made by methods known in the art and/or described in U.S. Pat. No. 6,696,038, U.S. App. Nos. 20030073619 and 20040142474 each of which is herein incorporated by reference in their entireties. The poly(alkylene imine) may be made using methods known in the art and/or as described in U.S. Pub. No. 20100004315, herein incorporated by reference in its entirety. The biodegradable polymer, biodegradable block copolymer, the biodegradable random copolymer, biodegradable polyester block copolymer, biodegradable polyester polymer, or biodegradable polyester random copolymer may be made using methods known in the art and/or as described in U.S. Pat. Nos. 6,517,869 and 6,267,987, the contents of which are each incorporated herein by reference in their entirety. The linear biodegradable copolymer may be made using methods known in the art and/or as described in U.S. Pat. No. 6,652,886. The PAGA polymer may be made using methods known in the art and/or as described in U.S. Pat. No. 6,217,912 herein incorporated by reference in its entirety. The PAGA polymer may be copolymerized to form a copolymer or block copolymer with polymers such as but not limited to, poly-L-lysine, polyargine, polyornithine, histones, avidin, protamines, polylactides and poly(lactide-co-glycolides). The biodegradable cross-linked cationic multi-block copolymers may be made my methods known in the art and/or as described in U.S. Pat. No. 8,057,821 or U.S. Pub. No. 2012009145 each of which are herein incorporated by reference in their entireties. For example, the multi-block copolymers may be synthesized using linear polyethyleneimine (LPEI) blocks which have distinct patterns as compared to branched polyethyleneimines. Further, the composition or pharmaceutical composition may be made by the methods known in the art, described herein, or as described in U.S. Pub. No. 20100004315 or U.S. Pat. Nos. 6,267,987 and 6,217,912 each of which are herein incorporated by reference in their entireties. [0266] The nucleic acid-based compositions of the disclosure may be formulated with at least one degradable polyester which may contain polycationic side chains. Degradable polyesters include, but are not limited to, poly(serine ester), poly(L-lactide-co-L-lysine), poly(4-hydroxy-L-proline ester), and combinations thereof. In some embodiments, the degradable polyesters may include a PEG conjugation to form a PEGylated polymer. [0267] The nucleic acid-based compositions of the disclosure may be formulated with at least one crosslinkable polyester. Crosslinkable polyesters include those known in the art and described in US Pub. No. 20120269761, herein incorporated by reference in its entirety. [0268] In some embodiments, the polymers described herein may be conjugated to a lipid- terminating PEG. As a non-limiting example, PLGA may be conjugated to a lipid-terminating PEG forming PLGA-DSPE-PEG. As another non-limiting example, PEG conjugates for use with the present disclosure are described in International Publication No. WO2008103276, herein incorporated by reference in its entirety. The polymers may be conjugated using a ligand conjugate such as, but not limited to, the conjugates described in U.S. Pat. No. 8,273,363, herein incorporated by reference in its entirety. [0269] In some embodiments, the nucleic acid-based compositions described herein may be conjugated with another compound. Non-limiting examples of conjugates are described in US Patent Nos. 7,964,578 and 7,833,992, each of which are herein incorporated by reference in their entireties. The nucleic acid-based compositions of the present disclosure may be conjugated with conjugates of formula 1-122 as described in US Patent Nos. 7,964,578 and 7,833,992, each of which are herein incorporated by reference in their entireties. The nucleic acid-based compositions described herein may be conjugated with a metal such as, but not limited to, gold. (See e.g., Giljohann et al. Journ. Amer. Chem. Soc. 2009131(6): 2072-2073; herein incorporated by reference in its entirety). In some embodiments, the nucleic acid-based compositions described herein may be conjugated and/or encapsulated in gold-nanoparticles. (International Pub. No. WO201216269 and U.S. Pub. No. 20120302940; each of which is herein incorporated by reference in its entirety). [0270] As described in U.S. Pub. No. 20100004313, herein incorporated by reference in its entirety, a gene delivery composition may include a nucleotide sequence and a poloxamer. For example, the nucleic acid-based compositions of the present disclosure may be used in a gene delivery composition with the poloxamer described in U.S. Pub. No. 20100004313. [0271] In some embodiments, the polymer formulation of the present disclosure may be stabilized by contacting the polymer formulation, which may include a cationic carrier, with a cationic lipopolymer which may be covalently linked to cholesterol and polyethylene glycol groups. The polymer formulation may be contacted with a cationic lipopolymer using the methods described in U.S. Pub. No. 20090042829 herein incorporated by reference in its entirety. [0272] The cationic carrier may include, but is not limited to, polyethylenimine, poly(trimethylenimine), poly(tetramethylenimine), polypropylenimine, aminoglycoside- polyamine, dideoxy-diamino-b-cyclodextrin, spermine, spermidine, poly(2- dimethylamino)ethyl methacrylate, poly(lysine), poly(histidine), poly(arginine), cationized gelatin, dendrimers, chitosan, 1,2-Dioleoyl-3-Trimethylammonium-Propane(DOTAP), N-[1- (2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA), 1-[2- (oleoyloxy)ethyl]-2-oleyl-3-(2-hydroxyethyl)imidazolinium chloride (DOTIM), 2,3- dioleyloxy-N-[2(sperminecarboxamido)ethyl]-N,N-dimethyl-1-propanaminium trifluoroacetate (DOSPA), 3B-[N—(N′,N′-Dimethylaminoethane)-carbamoyl]Cholesterol Hydrochloride (DC-Cholesterol HCl) diheptadecylamidoglycyl spermidine (DOGS), N,N- distearyl-N,N-dimethylammonium bromide (DDAB), N-(1,2-dimyristyloxyprop-3-yl)-N,N- dimethyl-N-hydroxyethyl ammonium bromide (DMRIE), N,N-dioleyl-N,N- dimethylammonium chloride DODAC) and combinations thereof. [0273] The nucleic acid-based compositions of the disclosure may be formulated in a polyplex of one or more polymers (U.S. Pub. No. 20120237565 and 20120270927; each of which is herein incorporated by reference in its entirety). In some embodiments, the polyplex comprises two or more cationic polymers. The cationic polymer may comprise a poly(ethylene imine) (PEI) such as linear PEI. [0274] The nucleic acid-based compositions of the disclosure can also be formulated as a nanoparticle using a combination of polymers, lipids, and/or other biodegradable agents, such as, but not limited to, calcium phosphate. Components may be combined in a core-shell, hybrid, and/or layer-by-layer architecture, to allow for fine-tuning of the nanoparticle so delivery of the nucleic acid-based compositions may be enhanced (Wang et al., Nat Mater. 20065:791-796; Fuller et al., Biomaterials.200829:1526-1532; DeKoker et al., Adv Drug Deliv Rev. 201163:748-761; Endres et al., Biomaterials. 201132:7721-7731; Su et al., Mol Pharm. 2011 Jun 6;8(3):774-87; herein incorporated by reference in its entirety). As a non- limiting example, the nanoparticle may comprise a plurality of polymers such as, but not limited to hydrophilic-hydrophobic polymers (e.g., PEG-PLGA), hydrophobic polymers (e.g., PEG) and/or hydrophilic polymers (International Pub. No. WO20120225129; herein incorporated by reference in its entirety). [0275] Biodegradable calcium phosphate nanoparticles in combination with lipids and/or polymers may be used to deliver nucleic acid-based compositions in vivo. In some embodiments, a lipid coated calcium phosphate nanoparticle, which may also contain a targeting ligand such as anisamide, may be used to deliver the nucleic acid-based compositions of the present disclosure. For example, to effectively deliver siRNA in a mouse metastatic lung model a lipid coated calcium phosphate nanoparticle was used (Li et al., J Contr Rel. 2010142: 416-421; Li et al., J Contr Rel. 2012158:108-114; Yang et al., Mol Ther. 201220:609-615; herein incorporated by reference in its entirety). This delivery system combines both a targeted nanoparticle and a component to enhance the endosomal escape, calcium phosphate, in order to improve delivery of the siRNA. [0276] In some embodiments, calcium phosphate with a PEG-polyanion block copolymer may be used to delivery nucleic acid-based compositions (Kazikawa et al., J Contr Rel. 2004 97:345-356; Kazikawa et al., J Contr Rel. 2006111:368-370; herein incorporated by reference in its entirety). [0277] In some embodiments, a PEG-charge-conversional polymer (Pitella et al., Biomaterials. 201132:3106-3114) may be used to form a nanoparticle to deliver the nucleic acid-based compositions of the present disclosure. The PEG-charge-conversional polymer may improve upon the PEG-polyanion block copolymers by being cleaved into a polycation at acidic pH, thus enhancing endosomal escape. [0278] The use of core-shell nanoparticles has additionally focused on a high-throughput approach to synthesize cationic cross-linked nanogel cores and various shells (Siegwart et al., Proc Natl Acad Sci U S A. 2011108:12996-13001). The complexation, delivery, and internalization of the polymeric nanoparticles can be precisely controlled by altering the chemical composition in both the core and shell components of the nanoparticle. For example, the core-shell nanoparticles may efficiently deliver nucleic acid-based compositions to mouse hepatocytes after they covalently attach cholesterol to the nanoparticle. [0279] In some embodiments, the nanoparticles described herein may be nanoparticles which include at least one ligand, and the ligand may be a peptide which targets tumors (i.e., a targeting peptide). As a non-limiting example, the targeting peptide is a cyclic RGD (cRGD) ligand which was found to bind to αvβ3 and αvβ5 integrins which are highly expressed on angiogenic endothelial cells of tumors as well as on a number of tumor cells. As a non-limiting example, the targeting peptide is an APRPG peptide (SEQ ID NO: 24457) which is an angiogenic vessel-homing peptide. As a non-limiting example, the targeting peptide is a cyclic NGR (CNGRCVSGCAGRC provided as SEQ ID NO: 24445) which is upregulated primarily in the tumor vasculature and in a few cancers such as fibrosarcomas. Linear NGR may also be an option for a targeting peptide. As a non-limiting example, the targeting peptide is a F3 peptide (KDEPQRRSARLSAKPAPPKPEPKPKKAPAKK provided as SEQ ID NO: 24446) which is 31-amino acid fragment of HMGN2 protein that binds to nucleolin which is often found on the cell surface oof tumor and angiogenic endothelial cells. As a non-limiting example, the targeting peptide is CGKRK (provided as SEQ ID NO: 24447) which may target the angiogenic vessels of tumors. As a non-limiting example, the targeting peptide is a tumor-penetrating peptide with the CendR Motif such as, but not limited to, LyP-1 (CGNKRTRGC, with the CendR motif underlined and the sequence provided as SEQ ID NO: 24448), iRGD (CRGDKGPDC, with the CendR motif underlined and the sequence provided as SEQ ID NO: 24449), and iNGR (CRNGRGPDC, with the CendR motif underlined and the sequence provided as SEQ ID NO: 24450). As a non- limiting example, the targeting peptide is a T7 peptide (HAIYPRH, provided as SEQ ID NO: 24451), a MMP2-cleavable peptide (GPLGIAGQ, provided as SEQ ID NO: 24452), a CP15 peptide (VHLGYAT, provided as SEQ ID NO: 24453), a FSH peptide (YTRDLVYKDPARPKIQKTCTF, provided as SEQ ID NO: 24454), gastrin-releasing peptides (GRPs) (CGGNHWAVGHLM, provided as SEQ ID NO: 24455), and a RVG-brain delivery peptide (YTIWMPENPRPGTPCDIFTNSRGKRASNG, provided as SEQ ID NO: 24456). As a non-limiting example, the ligand is a high molecular weight endogenous ligand such as transferrin (transferrin receptor is usually upregulated in many tumor cells), hyaluronic acid (HA) (negatively charged natural polymer which can bind to the surface CD44 receptors which are often overexpressed in primary and metastatic tumor cells), APOA1 (can bind with scavenger receptor class B-type 1 (SR-B1) which is found on the surface of hepatocytes). As a non-limiting example, the ligand is an aptamer, which is a small molecular weight (8-13 Kda) single-stranded RNA or DNA with low nanomolar binding affinities toward their targets. As a non-limiting example, the ligand is an antibody. As a non- limiting example, the ligand is a small molecule ligand such as, but not limited to, folate, anisamide, and galactose. (Leng et al. Journal of Drug Delivery. Vo. 17, Article ID 6971297; the contents of which is herein incorporated by reference in its entirety). [0280] In some embodiments, a hollow lipid core comprising a middle PLGA layer and an outer neutral lipid layer containing PEG may be used to delivery of the nucleic acid-based compositions of the present disclosure. As a non-limiting example, in mice bearing a luciferase-expressing tumor, it was determined that the lipid-polymer-lipid hybrid nanoparticle significantly suppressed luciferase expression, as compared to a conventional lipoplex (Shi et al, Angew Chem Int Ed. 201150:7027-7031; herein incorporated by reference in its entirety). [0281] In some embodiments, the lipid nanoparticles may comprise a core of the nucleic acid-based compositions disclosed herein and a polymer shell. The polymer shell may be any of the polymers described herein and are known in the art. In an additional embodiment, the polymer shell may be used to protect the modified nucleic acids in the core. [0282] Core–shell nanoparticles for use with the nucleic acid-based compositions of the present disclosure may be formed by the methods described in U.S. Pat. No. 8,313,777 herein incorporated by reference in its entirety. [0283] In some embodiments, the core-shell nanoparticles may comprise a core of the nucleic acid-based compositions disclosed herein and a polymer shell. The polymer shell may be any of the polymers described herein and are known in the art. In an additional embodiment, the polymer shell may be used to protect the nucleic acid-based compositions in the core. As a non-limiting example, the core-shell nanoparticle may be used to treat an eye disease or disorder (See e.g. US Publication No.20120321719, herein incorporated by reference in its entirety). [0284] In some embodiments, the polymer used with the formulations described herein may be a modified polymer (such as, but not limited to, a modified polyacetal) as described in International Publication No. WO2011120053, herein incorporated by reference in its entirety. Delivery [0285] The present disclosure encompasses the delivery of nucleic acid-based compositions including, for example, ASOs for any therapeutic, prophylactic, pharmaceutical, diagnostic or imaging use by any appropriate route taking into consideration likely advances in the sciences of drug delivery. Delivery may be naked or formulated. [0286] The nucleic acid-based compositions of the present disclosure may be delivered to a cell naked. As used herein in, “naked” refers to delivering nucleic acid-based compositions free from agents which promote transfection. For example, the nucleic acid-based compositions delivered to the cell may contain no modifications. The naked nucleic acid- based compositions may be delivered to the cell using routes of administration known in the art and described herein. [0287] The nucleic acid-based compositions of the present disclosure may be formulated, using the methods described herein. The formulations may contain nucleic acid-based compositions which may be modified and/or unmodified. The formulations may further include, but are not limited to, cell penetration agents, a pharmaceutically acceptable carrier, a delivery agent, a bioerodible or biocompatible polymer, a solvent, and a sustained-release delivery depot. The formulated nucleic acid-based compositions may be delivered to the cell using routes of administration known in the art and described herein. [0288] The compositions may also be formulated for direct delivery to an organ or tissue in any of several ways in the art including, but not limited to, direct soaking or bathing, via a catheter, by gels, powder, ointments, creams, gels, lotions, and/or drops, by using substrates such as fabric or biodegradable materials coated or impregnated with the compositions, and the like. The nucleic acid-based compositions of the present disclosure may also be cloned into a retroviral replicating vector (RRV) and transduced to cells. Administration [0289] The nucleic acid-based compositions of the present disclosure may be administered by any route which results in a therapeutically effective outcome. These include, but are not limited to enteral, gastroenteral, epidural, oral, transdermal, epidural (peridural), intracerebral (into the cerebrum), intracerebroventricular (into the cerebral ventricles), epicutaneous (application onto the skin), intradermal, (into the skin itself), subcutaneous (under the skin), nasal administration (through the nose), intravenous (into a vein), intraarterial (into an artery), intramuscular (into a muscle), intracardiac (into the heart), intraosseous infusion (into the bone marrow), intrathecal (into the spinal canal or into the subarachnoid space to reach the CSF), intraperitoneal, (infusion or injection into the peritoneum), intravesical infusion (e.g., into the bladder using a catheter), intravitreal, (through the eye), intracavernous injection, ( into the base of the penis), intravaginal administration, intrauterine, intraparenchymal (into the brain parenchyma), intracerebroventricular (into the cerebrospinal fluid), extra-amniotic administration, transdermal (diffusion through the intact skin for systemic distribution), transmucosal (diffusion through a mucous membrane), insufflation (snorting), sublingual, sublabial, enema, eye drops (onto the conjunctiva), in ear drops, nasal aerosol or inhalation. In specific embodiments, compositions may be administered in a way which allows them to cross the blood-brain barrier, vascular barrier, or other epithelial barrier. Routes of administration disclosed in International Publication WO 2013/090648 filed December 14, 2012, the contents of which are incorporated herein by reference in their entirety, may be used to administer the nucleic acid-based compositions of the present disclosure. [0290] Delivery of modified therapeutic compounds described herein to a subject over prolonged periods of time, for example, for periods of one week to one year, may be accomplished by a single administration of a controlled release system containing sufficient active ingredient for the desired release period. Various controlled release systems, such as monolithic or reservoir-type microcapsules, depot implants, polymeric hydrogels, osmotic pumps, vesicles, micelles, liposomes, transdermal patches, iontophoretic devices and alternative injectable dosage forms may be utilized for this purpose. Localization at the site to which delivery of the active ingredient is desired is an additional feature of some controlled release devices, which may prove beneficial in the treatment of certain disorders. [0291] In certain embodiments for transdermal administration, delivery across the barrier of the skin would be enhanced using electrodes (e.g. iontophoresis), electroporation, or the application of short, high-voltage electrical pulses to the skin, radiofrequencies, ultrasound (e.g. sonophoresis), microprojections (e.g. microneedles), jet injectors, thermal ablation, magnetophoresis, lasers, velocity, or photomechanical waves. The drug can be included in single-layer drug-in-adhesive, multi-layer drug-in-adhesive, reservoir, matrix, or vapor style patches, or could utilize patchless technology. Delivery across the barrier of the skin could also be enhanced using encapsulation, a skin lipid fluidizer, or a hollow or solid microstructured transdermal system (MTS, such as that manufactured by 3M), jet injectors. Additives to the formulation to aid in the passage of therapeutic compounds through the skin include prodrugs, chemicals, surfactants, cell penetrating peptides, permeation enhancers, encapsulation technologies, enzymes, enzyme inhibitors, gels, nanoparticles and peptide or protein chaperones. [0292] One form of controlled-release formulation contains the therapeutic compound or its salt dispersed or encapsulated in a slowly degrading, non-toxic, non-antigenic polymer such as copoly(lactic/glycolic) acid, as described in the pioneering work of Kent et al., US Patent No. 4,675,189, incorporated by reference herein. The compounds, or their salts, may also be formulated in cholesterol or other lipid matrix pellets, or silastomer matrix implants. Additional slow release, depot implant or injectable formulations will be apparent to the skilled artisan. See, for example, Sustained and Controlled Release Drug Delivery Systems, JR Robinson ed., Marcel Dekker Inc., New York, 1978; and Controlled Release of Biologically Active Agents, RW Baker, John Wiley & Sons, New York, 1987. The foregoing are incorporated by reference in their entirety. [0293] An additional form of controlled-release formulation comprises a solution of biodegradable polymer, such as copoly(lactic/glycolic acid) or block copolymers of lactic acid and PEG, is a bioacceptable solvent, which is injected subcutaneously or intramuscularly to achieve a depot formulation. Mixing of the therapeutic compounds described herein with such a polymeric formulation is suitable to achieve very long duration of action formulations. [0294] When formulated for nasal administration, the absorption across the nasal mucous membrane may be further enhanced by surfactants, such as, for example, glycocholic acid, cholic acid, taurocholic acid, ethocholic acid, deoxycholic acid, chenodeoxycholic acid, dehdryocholic acid, glycodeoxycholic acid, cycledextrins and the like in an amount in the range of between about 0.1 and 15 weight percent, between about 0.5 and 4 weight percent, or about 2 weight percent. An additional class of absorption enhancers reported to exhibit greater efficacy with decreased irritation is the class of alkyl maltosides, such as tetradecylmaltoside (Arnold, JJ et al., 2004, J Pharm Sci 93: 2205-13; Ahsan, F et al., 2001, Pharm Res 18:1742-46) and references therein, all of which are hereby incorporated by reference. [0295] The pharmaceutical compositions may be in the form of a sterile injectable preparation, for example, as a sterile injectable aqueous or oleaginous suspension. This suspension may be formulated according to techniques known in the art using suitable dispersing or wetting agents (such as, for example, Tween 80) and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non- toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are mannitol, water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil may be employed including synthetic mono- or diglycerides. Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions. These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant such as Ph. Helv or a similar alcohol. [0296] The pharmaceutical compositions of the present disclosure may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, and aqueous suspensions and solutions. In the case of tablets for oral use, carriers that are commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried corn starch. When aqueous suspensions are administered orally, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening and/or flavoring and/or coloring agents may be added. [0297] The pharmaceutical compositions of present disclosure may also be administered in the form of suppositories for rectal administration. These compositions can be prepared by mixing a compound of the present disclosure with a suitable non-irritating excipient that is solid at room temperature but liquid at the rectal temperature and therefore will melt in the rectum to release the active components. Such materials include, but are not limited to, cocoa butter, beeswax and polyethylene glycols. [0298] Topical administration of the pharmaceutical compositions of the present disclosure is especially useful when the desired treatment involves areas or organs readily accessible by topical application. For application topically to the skin, the pharmaceutical composition should be formulated with a suitable ointment containing the active components suspended or dissolved in a carrier. Carriers for topical administration of the compounds of the present disclosure include, but are not limited to, mineral oil, liquid petroleum, white petroleum, propylene glycol, polyoxyethylene polyoxypropylene compound, emulsifying wax and water. Alternatively, the pharmaceutical composition can be formulated with a suitable lotion or cream containing the active compound suspended or dissolved in a carrier. Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water. The pharmaceutical compositions of the present disclosure may also be topically applied to the lower intestinal tract by rectal suppository formulation or in a suitable enema formulation. Topical transdermal patches are also included in the present disclosure. [0299] The pharmaceutical compositions of the present disclosure may be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well- known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other solubilizing or dispersing agents known in the art. [0300] When formulated for delivery by inhalation, a number of formulations offer advantages. Adsorption of the therapeutic compound to readily dispersed solids such as diketopiperazines (for example, Technosphere particles (Pfutzner, A and Forst, T, 2005, Expert Opin Drug Deliv 2:1097-1106) or similar structures gives a formulation that results in rapid initial uptake of the therapeutic compound. Lyophilized powders, especially glassy particles, containing the therapeutic compound and an excipient are useful for delivery to the lung with good bioavailability, for example, see Exubera® (inhaled insulin, Pfizer, Inc. and Aventis Pharmaceuticals Inc.) and Afrezza® (inhaled insulin, Mannkind, Corp.). [0301] The pharmaceutical compositions of the present disclosure may be administered by local delivery to the bladder such as, but not limited to, intravesical therapy. Such compositions are prepared according to techniques well-known in the art of pharmaceutical formulation and may be prepared as solutions that can be delivered using, for example, a catheter that is put into the bladder through the urethra. [0302] The pharmaceutical compositions of the present disclosure may be formulated to be administered to the CNS by routes known in the art such as, but not limited to, direct intraparenchymal administration, intrathecal delivery and intracerebroventricular infusion. In some embodiments, the pharmaceutical compositions are formulated to have the biodistribution of the pharmaceutical composition located in the tumor cells. [0303] In some embodiments, the pharmaceutical compositions of the present disclosure may be formulated to improve delivery to tumors. Dosage Forms [0304] A pharmaceutical composition described herein can be formulated into a dosage form described herein, such as a topical, intranasal, intratracheal, or injectable (e.g., intravenous, intraocular, intravitreal, intramuscular, intracardiac, intraperitoneal, subcutaneous). Liquid dosage forms, injectable preparations, pulmonary forms, and solid dosage forms described in International Publication WO 2013/090648 filed December 14, 2012, the contents of which are incorporated herein by reference in their entirety may be used as dosage forms for the nucleic acid-based compositions of the present disclosure. [0305] The present disclosure contemplates dosage levels of between about 0.001 and about 100 mg GABPB1-hybridizing oligonucleotide (e.g., GABPB1-targeting ASO)/kg body weight per day, preferably between about 0.005 and about 50 mg/kg, 0.01 and about 10 mg/kg, 0.05 and about 5 mg/kg, 0.1 and about 1 mg/kg body weight. Other embodiments contemplate a dosage of between about 0.001-0.010, 0.010-0.050, 0.050-0.100, 0.1-0.5, 0.5- 1.0, 1.0-5.0, 5.0-10, or 10-50 mg/kg body weight. The dosages may be administered about hourly, multiple times per day, daily, every other day, weekly, every other week, monthly, or on an as-needed basis. [0306] Such administration can be used as a chronic or acute therapy. The amount of drug that may be combined with the carrier to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. A typical preparation will contain from about 5% to about 95% active compound (w/w). Preferably, such preparations contain from about 20% to about 80%, 30% to about 70%, 40% to about 60%, or about 50% active compound. In other embodiments, the preparations used in the present disclosure will be about 5-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90- 99%, or greater than 99% of the active ingredient. [0307] Upon improvement of a patient's condition, a maintenance dose of a compound, composition or combination of the present disclosure may be administered, if necessary. Subsequently, the dosage or frequency of administration, or both, may be reduced, as a function of the symptoms, to a level at which the improved condition is retained when the symptoms have been alleviated to the desired level, treatment should cease. Patients may, however, require intermittent treatment on a long-term basis upon any recurrence of disease symptoms. [0308] As the skilled artisan will appreciate, lower or higher doses than those recited above may be required. Specific dosage and treatment regimens for any particular patient will depend upon a variety of factors, including the activity of the specific compound employed, the age, body weight, general health status, gender, diet, time of administration, rate of excretion, drug combination, the severity and course of an infection, the patient's disposition to the infection and the judgment of the treating physician. III. Methods of Use [0309] One aspect of the present disclosure provides methods of using nucleic acid-based compositions of the present disclosure and pharmaceutical compositions comprising the nucleic acid-based compositions and at least one pharmaceutically acceptable carrier. The nucleic acid-based compositions of the present disclosure modulate the expression of the target gene, i.e., GABPB1. In some embodiments is provided a method of regulating the expression of a target gene in vitro and/or in vivo comprising administering the nucleic acid- based compositions of the present disclosure. In some embodiments, the expression of the target gene is decreased by at least 5%, 10%, 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the nucleic acid-based compositions of the present disclosure compared to the expression of the target gene in the absence of the nucleic acid-based compositions of the present disclosure. In further embodiments, the expression of the target gene is decreased by a factor of at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or by a factor of at least 15, 20, 25, 30, 35, 40, 45, 50, or by a factor of at least 60, 70, 80, 90, 100, in the presence of the nucleic acid-based compositions of the present disclosure compared to the expression of the target gene in the absence of the nucleic acid-based compositions of the present disclosure. [0310] One aspect of the present application provides a method of modulating the expression of GABPB1 gene comprising administering GABPB1-hybridizing oligonucleotides of the present disclosure. In some embodiments, the expression of GABPB1 gene is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure compared to the expression of GABPB1 gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure. In some embodiments, the expression of GABPB1 gene is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure which specifically targets the GABPB1L isoform as compared to the expression of GABPB1 gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure specifically targeting GABPB1L. In some embodiments, the expression of GABPB1 gene is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1- hybridizing oligonucleotides of the present disclosure which specifically targets the GABPB1S isoform as compared to the expression of GABPB1 gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure specifically targeting GABPB1S. In some embodiments, the expression of GABPB1 gene is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure which specifically targets both the GABPB1L and GABPB1S isoforms (total GABPB1) as compared to the expression of GABPB1 gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure specifically targeting total GABPB1. In further embodiments, the expression of GABPB1 gene is decreased by a factor of at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or by a factor of at least 15, 20, 25, 30, 35, 40, 45, 50, or by a factor of at least 60, 70, 80, 90, 100, in the presence of the GABPB1-hybridizing oligonucleotides (e.g., GABPB1-hybridizing oligonucleotides targeting the GABPB1L isoform, the GABPB1S isoform or total GABPB1) of the present disclosure compared to the expression of GABPB1 gene in the absence of the GABP-hybridizing oligonucleotides of the present disclosure. [0311] In some embodiments, the expression of TERT is decreased by at least 20%, 30%, 40%, or at least 45%, 50%, 55%, 60%, 65%, 70%, 75%, or at least 80% in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure compared to the expression of TERT in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure. In further embodiments, the expression of TERT gene is decreased by a factor of at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or by a factor of at least 15, 20, 25, 30, 35, 40, 45, 50, or by a factor of at least 60, 70, 80, 90, 100, in the presence of the GABPB1-hybridizing oligonucleotides of the present disclosure compared to the expression of TERT gene in the absence of the GABPB1-hybridizing oligonucleotides of the present disclosure. The modulation of the expression of GABPB1 gene may be reflected in or determined by a change in TERT protein levels. [0312] Some embodiments provide methods of use of the nucleic acid-based compositions described herein to prevent or treat diseases or disorders associated with telomerase expression such as, but not limited to cancer. Thus, in some embodiments, the methods provided herein include administering GABPB1-hybridizing oligonucleotides described herein to subjects having a cancer. The methods of administering the GABPB1-hybridizing oligonucleotides decrease expression of the GABPB1 gene, and, in some cases, decrease the level of TERT mRNA expression in cancer cells harboring TERT promoter mutations which may ultimately reduce the expression of telomerase in the subject. Accordingly, the present disclosure provides methods for treating individual subjects suffering from cancer. [0313] In some embodiments, the methods of use can be assessed using any endpoint indicating a benefit to the subject, including, without limitation, (1) inhibition, to some extent, of disease progression, including stabilization, slowing down and complete arrest; (2) reduction in the number of disease episodes and/or symptoms; (3) inhibition (i.e., reduction, slowing down or complete stopping) of a disease cell infiltration into adjacent peripheral organs and/or tissues; (4) inhibition (i.e. reduction, slowing down or complete stopping) of disease spread; (5) decrease of an autoimmune condition; (6) favorable change in the expression of a biomarker associated with the disorder; (7) relief, to some extent, of one or more symptoms associated with a disorder; (8) increase in the length of disease-free presentation following treatment; or (9) decreased mortality at a given point of time following treatment Cancer [0314] Various cancers may be treated with GABPB1-hybridizing oligonucleotides described herein. As used herein, the term “cancer” refers to any of various malignant neoplasms characterized by the proliferation of anaplastic cells that tend to invade surrounding tissue and metastasize to new body sites and also refers to the pathological condition characterized by such malignant neoplastic growths. Cancers may be tumors or hematological malignancies, and include but are not limited to, all types of lymphomas/leukemias, carcinomas and sarcomas, such as those cancers or tumors found in the anus, bladder, bile duct, bone, brain, breast, cervix, colon/rectum, endometrium, esophagus, eye, gallbladder, head and neck, liver, kidney, larynx, lung, mediastinum (chest), mouth, ovaries, pancreas, penis, prostate, skin, small intestine, stomach, spinal marrow, tailbone, testicles, thyroid and uterus. [0315] Types of carcinomas which may be treated with the compositions of the present disclosure include, but are not limited to, papilloma/carcinoma, choriocarcinoma, endodermal sinus tumor, teratoma, adenoma/adenocarcinoma, melanoma, fibroma, lipoma, leiomyoma, rhabdomyoma, mesothelioma, angioma, osteoma, chondroma, glioma, lymphoma/leukemia, squamous cell carcinoma, small cell carcinoma, large cell undifferentiated carcinomas, basal cell carcinoma and sinonasal undifferentiated carcinoma. [0316] Types of carcinomas which may be treated with the compositions of the present disclosure include, but are not limited to, soft tissue sarcoma such as alveolar soft part sarcoma, angiosarcoma, dermatofibrosarcoma, desmoid tumor, desmoplastic small round cell tumor, extraskeletal chondrosarcoma, extraskeletal osteosarcoma, fibrosarcoma, hemangiopericytoma, hemangiosarcoma, Kaposi's sarcoma, leiomyosarcoma, liposarcoma, lymphangiosarcoma, lymphosarcoma, malignant fibrous histiocytoma, neurofibrosarcoma, rhabdomyosarcoma, synovial sarcoma, and Askin's tumor, Ewing's sarcoma (primitive neuroectodermal tumor), malignant hemangioendothelioma, malignant schwannoma, osteosarcoma, and chondrosarcoma. [0317] As a non-limiting example, the carcinoma which may be treated may be Acute granulocytic leukemia, Acute lymphocytic leukemia, Acute myelogenous leukemia, Adenocarcinoma, Adenosarcoma, Adrenal cancer, Adrenocortical carcinoma, Anal cancer, Anaplastic astrocytoma, Angiosarcoma, Appendix cancer, Astrocytoma, Basal cell carcinoma, B-Cell lymphoma ), Bile duct cancer, Bladder cancer, Bone cancer, Bowel cancer, Brain cancer, Brain stem glioma, Brain tumor, Breast cancer, Carcinoid tumors, Cervical cancer, Cholangiocarcinoma, Chondrosarcoma, Chronic lymphocytic leukemia, Chronic myelogenous leukemia, Colon cancer, Colorectal cancer, Craniopharyngioma, Cutaneous lymphoma, Cutaneous melanoma, Diffuse astrocytoma, Ductal carcinoma in situ, Endometrial cancer, Ependymoma, Epithelioid sarcoma, Esophageal cancer, Ewing sarcoma, Extrahepatic bile duct cancer, Eye cancer, Fallopian tube cancer, Fibrosarcoma, Gallbladder cancer, Gastric cancer, Gastrointestinal cancer, Gastrointestinal carcinoid cancer, Gastrointestinal stromal tumors, General, Germ cell tumor, Glioblastoma multiforme, Glioma, Hairy cell leukemia, Head and neck cancer, Hemangioendothelioma, Hodgkin lymphoma, Hodgkin's disease, Hodgkin's lymphoma, Hypopharyngeal cancer, Infiltrating ductal carcinoma, Infiltrating lobular carcinoma, Inflammatory breast cancer, Intestinal Cancer, Intrahepatic bile duct cancer, Invasive / infiltrating breast cancer, Islet cell cancer, Jaw cancer, Kaposi sarcoma, Kidney cancer, Laryngeal cancer, Leiomyosarcoma, Leptomeningeal metastases, Leukemia, Lip cancer, Liposarcoma, Liver cancer, Lobular carcinoma in situ, Low-grade astrocytoma, Lung cancer, Lymph node cancer, Lymphoma, Male breast cancer, Medullary carcinoma, Medulloblastoma, Melanoma, Meningioma, Merkel cell carcinoma, Mesenchymal chondrosarcoma, Mesenchymous, Mesothelioma, Metastatic breast cancer, Metastatic melanoma, Metastatic squamous neck cancer, Mixed gliomas, Mouth cancer, Mucinous carcinoma, Mucosal melanoma, Multiple myeloma, Nasal cavity cancer, Nasopharyngeal cancer, Neck cancer, Neuroblastoma, Neuroendocrine tumors, Non-Hodgkin lymphoma, Non-Hodgkin's lymphoma, Non-small cell lung cancer, Oat cell cancer, Ocular cancer, Ocular melanoma, Oligodendroglioma, Oral cancer, Oral cavity cancer, Oropharyngeal cancer, Osteogenic sarcoma, Osteosarcoma, Ovarian cancer, Ovarian epithelial cancer, Ovarian germ cell tumor, Ovarian primary peritoneal carcinoma, Ovarian sex cord stromal tumor, Paget's disease, Pancreatic cancer, Papillary carcinoma, Paranasal sinus cancer, Parathyroid cancer, Pelvic cancer, Penile cancer, Peripheral nerve cancer, Peritoneal cancer, Pharyngeal cancer, Pheochromocytoma, Pilocytic astrocytoma, Pineal region tumor, Pineoblastoma, Pituitary gland cancer, Primary central nervous system lymphoma, Prostate cancer, Rectal cancer, Renal cell cancer, Renal pelvis cancer, Rhabdomyosarcoma, Salivary gland cancer, Sarcoma, Sarcoma, bone, Sarcoma, soft tissue, Sarcoma, uterine, Sinus cancer, Skin cancer, Small cell lung cancer, Small intestine cancer, Soft tissue sarcoma, Spinal cancer, Spinal column cancer, Spinal cord cancer, Spinal tumor, Squamous cell carcinoma, Stomach cancer, Synovial sarcoma, T-cell lymphoma ), Testicular cancer, Throat cancer, Thymoma / thymic carcinoma, Thyroid cancer, Tongue cancer, Tonsil cancer, Transitional cell cancer, Transitional cell cancer, Transitional cell cancer, Triple- negative breast cancer, Tubal cancer, Tubular carcinoma, Ureteral cancer, Ureteral cancer, Urethral cancer, Uterine adenocarcinoma, Uterine cancer, Uterine sarcoma, Vaginal cancer, and Vulvar cancer. IV. Kits and Devices Kits [0318] The disclosure provides a variety of kits for conveniently and/or effectively carrying out methods of the present disclosure. Typically, kits will comprise sufficient amounts and/or numbers of components to allow a user to perform multiple treatments of a subject(s) and/or to perform multiple experiments. [0319] In some embodiments, the present disclosure provides kits for modulating the expression of genes in vitro or in vivo, comprising nucleic acid-based compositions of the present disclosure or a combination of nucleic acid-based compositions of the present disclosure, nucleic acid-based compositions modulating other genes, siRNAs, miRNAs or other oligonucleotide molecules. [0320] The kit may further comprise packaging and instructions and/or a delivery agent to form a formulation, e.g., for administration to a subject in need of treatment using the nucleic acid-based compositions described herein. The delivery agent may comprise a saline, a buffered solution, a lipidoid, a dendrimer or any suitable delivery agent. [0321] In one non-limiting example, the buffer solution may include sodium chloride, calcium chloride, phosphate and/or EDTA. In another non-limiting example, the buffer solution may include, but is not limited to, saline, saline with 2mM calcium, 5% sucrose, 5% sucrose with 2mM calcium, 5% Mannitol, 5% Mannitol with 2mM calcium, Ringer’s lactate, sodium chloride, sodium chloride with 2mM calcium and mannose (See U.S. Pub. No. 20120258046; herein incorporated by reference in its entirety). In yet another non-limiting example, the buffer solutions may be precipitated or it may be lyophilized. The amount of each component may be varied to enable consistent, reproducible higher concentration saline or simple buffer formulations. The components may also be varied in order to increase the stability of nucleic acid-based compositions in the buffer solution over a period of time and/or under a variety of conditions. Devices [0322] The present disclosure provides for devices which may incorporate nucleic acid- based compositions of the present disclosure. These devices can contain a stable formulation available to be immediately delivered to a subject in need thereof, such as a human patient. [0323] Non-limiting examples of the devices include a pump, a catheter, a needle, a transdermal patch, a pressurized olfactory delivery device, electroporation devices, iontophoresis devices, multi-layered microfluidic devices. The devices may be employed to deliver nucleic acid-based compositions of the present disclosure according to single, multi- or split-dosing regiments. The devices may be employed to deliver nucleic acid-based compositions of the present disclosure across biological tissue, intradermal, subcutaneously, or intramuscularly. More examples of devices suitable for delivering oligonucleotides are disclosed in International Publication WO 2013/090648, the contents of which are incorporated herein by reference in their entirety. V. Definitions [0324] For convenience, the meaning of certain terms and phrases used in the specification, examples, and appended claims, are provided below. If there is an apparent discrepancy between the usage of a term in other parts of this specification and its definition provided in this section, the definition in this section shall prevail. [0325] About: As used herein, the term “about” means +/- 10% of the recited value. [0326] Administered in combination: As used herein, the term “administered in combination” or “combined administration” means that two or more agents are administered to a subject at the same time or within an interval such that there may be an overlap of an effect of each agent on the patient. In some embodiments, they are administered within about 60, 30, 15, 10, 5, or 1 minute of one another. In some embodiments, the administrations of the agents are spaced sufficiently close together such that a combinatorial (e.g., a synergistic) effect is achieved. [0327] Agonist: As used herein, the term “agonist” refers to a substance that binds to a receptor and activates the receptor to produce a biological response. They can be in the form of antibodies, antigen-binding fragments, proteins, peptides, glycoproteins, glycopeptides, glycolipids, polysaccharides, oligosaccharides, nucleic acids, bioorganic molecules, peptidomimetics, chemicals, pharmacological agents and their metabolites, and the like. In contrast, an "antagonist" refers to a molecule capable of neutralizing, blocking, inhibiting, abrogating, reducing or interfering with the activities of a particular or specified protein, including its binding to one or more receptors in the case of a ligand, or binding to one or more ligands in case of a receptor. [0328] Amino acid: As used herein, the terms "amino acid" and "amino acids" refer to all naturally occurring L-alpha-amino acids. The amino acids are identified by either the one- letter or three-letter designations as follows: aspartic acid (Asp:D), isoleucine (Ile:I), threonine (Thr:T), leucine (Leu:L), serine (Ser:S), tyrosine (Tyr:Y), glutamic acid (Glu:E), phenylalanine (Phe:F), proline (Pro:P), histidine (His:H), glycine (Gly:G), lysine (Lys:K), alanine (Ala:A), arginine (Arg:R), cysteine (Cys:C), tryptophan (Trp:W), valine (Val:V), glutamine (Gln:Q) methionine (Met:M), asparagines (Asn:N), where the amino acid is listed first followed parenthetically by the three and one letter codes, respectively. [0329] Animal: As used herein, the term “animal” refers to any member of the animal kingdom. In some embodiments, “animal” refers to humans at any stage of development. In some embodiments, “animal” refers to non-human animals at any stage of development. In certain embodiments, the non-human animal is a mammal (e.g., a rodent, a mouse, a rat, a rabbit, a monkey, a dog, a cat, a sheep, cattle, a primate, or a pig). In some embodiments, animals include, but are not limited to, mammals, birds, reptiles, amphibians, fish, and worms. In some embodiments, the animal is a transgenic animal, genetically-engineered animal, or a clone. [0330] Antisense oligonucleotide (ASO): “Antisense oligonucleotides” or “ASOs” are typically short (e.g., around 8 – 50 nucleotides), single-stranded oligonucleotide molecules designed to hybridize with an RNA sequence, e.g., mRNA, rRNA. Such targeting may result in either an upregulation or downregulation of the expression of the gene depending on the design of the ASO and the site of action, or localization of the ASO. In the context of the present disclosure, ASOs include oligodeoxyribonucleotide or oligoribonucleotide molecules that downregulate or have a negative effect on the expression of a specific gene. The ASOs can be between 8 to 20 nucleotides in length. The ASOs are designed to complementarily bind to a region of a target gene RNA, including pre-mRNA or mRNA, the target gene being GABPB1. For example, an ASO that downregulates the expression of the GABPB1 gene is called a “GABPB1-ASO.” ASOs as described herein include ASOs having modified nucleotides, including fully modified and partially modified ASOs. “Gapmer ASOs” are short single- stranded ASOs containing a central DNA sequence commonly flanked by a locked nucleic acid (LNA) sequence that interrupts mRNA expression by induction of RNase H activation. They can exhibit cellular entry without the necessity of a transfection agent by a process termed gymnosis. [0331] Approximately: As used herein, the term “approximately” or “about,” as applied to one or more values of interest, refers to a value that is similar to a stated reference value. In certain embodiments, the term “approximately” or “about” refers to a range of values that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in either direction (greater than or less than) of the stated reference value unless otherwise stated or otherwise evident from the context (except where such number would exceed 100% of a possible value). [0332] Associated with: As used herein, the terms “associated with,” “conjugated,” “linked,” “attached,” and “tethered,” when used with respect to two or more moieties, means that the moieties are physically associated or connected with one another, either directly or via one or more additional moieties that serves as a linking agent, to form a structure that is sufficiently stable so that the moieties remain physically associated under the conditions in which the structure is used, e.g., physiological conditions. An “association” need not be strictly through direct covalent chemical bonding. It may also suggest ionic or hydrogen bonding or a hybridization based connectivity sufficiently stable such that the “associated” entities remain physically associated. [0333] Bifunction or Bifunctional: As used herein, the terms “bifunction” and “bifunctional” refers to any substance, molecule or moiety which is capable of or maintains at least two functions. The functions may affect the same outcome or a different outcome. The structure that produces the function may be the same or different. For example, bifunctional nucleic acid-based compositions of the present disclosure may comprise a cytotoxic peptide (a first function) while those nucleosides which comprise the nucleic acid-based compositions are, in and of themselves, cytotoxic (second function). [0334] Biocompatible: As used herein, the term “biocompatible” means compatible with living cells, tissues, organs or systems posing little to no risk of injury, toxicity or rejection by the immune system. [0335] Biodegradable: As used herein, the term “biodegradable” means capable of being broken down into innocuous products by the action of living things. [0336] Biologically active: As used herein, the phrase “biologically active” refers to a characteristic of any substance that has activity in a biological system and/or organism. For instance, a substance that, when administered to an organism, has a biological effect on that organism, is considered to be biologically active. In particular embodiments, the nucleic acid- based compositions of the present disclosure may be considered biologically active if even a portion of the nucleic acid-based compositions is biologically active or mimics an activity considered biologically relevant. [0337] Cancer: As used herein, the term "cancer" in an individual refers to the presence of cells possessing characteristics typical of cancer-causing cells, such as uncontrolled proliferation, immortality, metastatic potential, rapid growth and proliferation rate, and certain characteristic morphological features. Often, cancer cells will be in the form of a tumor, but such cells may exist alone within an individual, or may circulate in the blood stream as independent cells, such as leukemic cells. [0338] Cell growth: As used herein, the term “cell growth” is principally associated with growth in cell numbers, which occurs by means of cell reproduction (i.e. proliferation) when the rate of the latter is greater than the rate of cell death (e.g. by apoptosis or necrosis), to produce an increase in the size of a population of cells, although a small component of that growth may in certain circumstances be due also to an increase in cell size or cytoplasmic volume of individual cells. An agent that inhibits cell growth can thus do so by either inhibiting proliferation or stimulating cell death, or both, such that the equilibrium between these two opposing processes is altered. [0339] Cell type: As used herein, the term "cell type" refers to a cell from a given source (e.g., a tissue, organ) or a cell in a given state of differentiation, or a cell associated with a given pathology or genetic makeup. [0340] Chromosome: As used herein, the term “chromosome” refers to an organized structure of DNA and protein found in cells. [0341] Complementary: As used herein, the term “complementary” as it relates to nucleic acids refers to hybridization or base pairing between nucleotides or nucleic acids, such as, for example, between the two strands of a double-stranded DNA molecule or between an oligonucleotide probe and a target. [0342] Condition: As used herein, the term “condition” refers to the status of any cell, organ, organ system or organism. Conditions may reflect a disease state or simply the physiologic presentation or situation of an entity. Conditions may be characterized as phenotypic conditions such as the macroscopic presentation of a disease or genotypic conditions such as the underlying gene or protein expression profiles associated with the condition. Conditions may be benign or malignant. [0343] Controlled Release: As used herein, the term “controlled release” refers to a pharmaceutical composition or compound release profile that conforms to a particular pattern of release to effect a therapeutic outcome. [0344] Cytotoxic: As used herein, “cytotoxic” refers to killing or causing injurious, toxic, or deadly effect on a cell (e.g., a mammalian cell (e.g., a human cell)), bacterium, virus, fungus, protozoan, parasite, prion, or a combination thereof. [0345] Delivery: As used herein, “delivery” refers to the act or manner of delivering a compound, substance, entity, moiety, cargo or payload. [0346] Delivery Agent: As used herein, “delivery agent” refers to any substance which facilitates, at least in part, the in vivo delivery of nucleic acid-based compositions of the present disclosure to targeted cells. [0347] Destabilized: As used herein, the term “destable,” “destabilize,” or “destabilizing region” means a region or molecule that is less stable than a starting, wild-type or native form of the same region or molecule. [0348] Detectable label: As used herein, “detectable label” refers to one or more markers, signals, or moieties which are attached, incorporated or associated with another entity that is readily detected by methods known in the art including radiography, fluorescence, chemiluminescence, enzymatic activity, absorbance and the like. Detectable labels include radioisotopes, fluorophores, chromophores, enzymes, dyes, metal ions, ligands such as biotin, avidin, streptavidin and haptens, quantum dots, and the like. Detectable labels may be located at any position in the oligonucleotides disclosed herein. They may be within the nucleotides or located at the 5’ or 3’ terminus. [0349] Downregulation: The terms “downregulation” or “inactivation” as used in the context of a gene refer to a decrease in the level of expression of the gene, a decrease in the level of the polypeptide(s) encoded by the gene, the level bioactive gene product, or level of the RNA transcript(s) transcribed from the template strand of the gene as compared to the levels observed in the absence of the nucleic acid-based composition(s) described herein. The nucleic acid-based compositions of the present disclosure may have a direct downregulating effect on the expression of the target gene. The term “downregulation,” depending on the context, can also refer to the effect of a trans-acting activator of expression. [0350] Downregulatory sequence element: The terms “DSE” or “downregulatory sequence element” as used in the context of expression control elements in a gene or a transcript of a gene, including a target transcript, refer to those cis-acting sequence-based or secondary structure-based features of a sequence which, under normal physiological conditions, tend to decrease or inhibit translation of the transcript and/or expression of the gene. The term “downregulation,” depending on the context, can also refer to the effect of a trans-acting inhibitor of expression. [0351] Effective Amount: As used herein, an "effective amount" refers to an amount of therapeutic compound that is effective, at dosages and for periods of time necessary, to achieve the desired therapeutic or prophylactic result. [0352] Element: The term “element” as used herein in connection with a region or feature of a nucleotide sequence refers to a particular feature of the sequence, such as a known binding site for a protein or other factor or a region that is involved in the formation of secondary structure of the nucleotide sequence. Elements include, for example, upstream open reading frames (uORFs), internal ribosome entry sites (IRES), upstream initiation codons, upstream termination codons, and binding sites for proteins and other factors that operate in trans with respect to the nucleotide sequence. [0353] Encapsulate: As used herein, the term “encapsulate” means to enclose, surround or encase. [0354] Engineered: As used herein, embodiments of the disclosure are “engineered” when they are designed to have a feature or property, whether structural or chemical, that varies from a starting point, wild type or native molecule. [0355] Equivalent subject: As used herein, “equivalent subject" may be e.g. a subject of similar age, sex and health such as liver health or cancer stage, or the same subject prior to treatment according to the disclosure. The equivalent subject is "untreated" in that he does not receive treatment with nucleic acid-based compositions according to the disclosure. However, he may receive a conventional anti-cancer treatment, provided that the subject who is treated with the nucleic acid-based compositions of the disclosure receives the same or equivalent conventional anti-cancer treatment. [0356] Exosome: As used herein, “exosome” is a vesicle secreted by mammalian cells. [0357] Expression: As used herein, “expression” of a nucleic acid sequence refers to one or more of the following events: (1) production of an RNA template from a DNA sequence (e.g., by transcription); (2) processing of an RNA transcript (e.g., by splicing, editing, 5’ cap formation, and/or 3’ end processing); (3) translation of an RNA into a polypeptide or protein; and (4) post-translational modification of a polypeptide or protein. [0358] Feature: As used herein, a “feature” refers to a characteristic, a property, or a distinctive element. [0359] Formulation: As used herein, a “formulation” includes at least one nucleic acid- based composition of the present disclosure and a delivery agent. [0360] Fragment: A “fragment,” as used herein, refers to a portion. For example, fragments of proteins may comprise polypeptides obtained by digesting full-length protein isolated from cultured cells. Fragments of oligonucleotides may comprise nucleotides, or regions of nucleotides. [0361] Functional: As used herein, a “functional” biological molecule is a biological molecule in a form in which it exhibits a property and/or activity by which it is characterized. [0362] Gene: As used herein, the term "gene" refers to a nucleic acid sequence that comprises control and most often coding sequences necessary for producing a polypeptide or precursor. Genes, however, may not be translated and instead code for regulatory or structural RNA molecules. [0363] A gene may be derived in whole or in part from any source known to the art, including a plant, a fungus, an animal, a bacterial genome or episome, eukaryotic, nuclear or plasmid DNA, cDNA, viral DNA, or chemically synthesized DNA. A gene may contain one or more modifications in either the coding or the untranslated regions that could affect the biological activity or the chemical structure of the expression product, the rate of expression, or the manner of expression control. Such modifications include, but are not limited to, mutations, insertions, deletions, and substitutions of one or more nucleotides. The gene may constitute an uninterrupted coding sequence or it may include one or more introns, bound by the appropriate splice junctions. [0364] Gene expression: As used herein, the term "gene expression" refers to the process by which a nucleic acid sequence undergoes successful transcription and in most instances translation to produce a protein or peptide. For clarity, when reference is made to measurement of “gene expression”, this should be understood to mean that measurements may be of the nucleic acid product of transcription, e.g., RNA or mRNA or of the amino acid product of translation, e.g., polypeptides or peptides. Methods of measuring the amount or levels of RNA, mRNA, polypeptides and peptides are well known in the art. [0365] Genome: The term "genome" is intended to include the entire DNA complement of an organism, including the nuclear DNA component, chromosomal or extrachromosomal DNA, as well as the cytoplasmic domain (e.g., mitochondrial DNA). [0366] Homolog: As used herein, the term “homologs” are bioactive molecules that are similar to a reference molecule at the nucleotide sequence, peptide sequence, functional, or structural level. Homologs may include sequence derivatives that share a certain percent identity with the reference sequence. Homologous or derivative nucleic acid sequences may also be defined by their ability to remain bound to a reference nucleic acid sequence under high stringency hybridization conditions. Homologs having a structural or functional similarity to a reference molecule may be chemical derivatives of the reference molecule. [0367] Homology: As used herein, the term “homology” refers to the overall relatedness between polymeric molecules, e.g. between nucleic acid molecules (e.g. DNA molecules and/or RNA molecules) and/or between polypeptide molecules. In some embodiments, polymeric molecules are considered to be “homologous” to one another if their sequences are at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical or similar. The term “homologous” necessarily refers to a comparison between at least two sequences (polynucleotide or polypeptide sequences). In accordance with the disclosure, two polynucleotide sequences are considered to be homologous if the polypeptides they encode are at least about 50%, 60%, 70%, 80%, 90%, 95%, or even 99% for at least one stretch of at least about 20 amino acids. In some embodiments, homologous polynucleotide sequences are characterized by the ability to encode a stretch of at least 4–5 uniquely specified amino acids. For polynucleotide sequences less than 60 nucleotides in length, homology is determined by the ability to encode a stretch of at least 4–5 uniquely specified amino acids. In accordance with the disclosure, two protein sequences are considered to be homologous if the proteins are at least about 50%, 60%, 70%, 80%, or 90% identical for at least one stretch of at least about 20 amino acids. [0368] Identity: As used herein, the term “identity” refers to the overall relatedness between polymeric molecules, e.g., between oligonucleotide molecules (e.g. DNA molecules and/or RNA molecules) and/or between polypeptide molecules. Calculation of the percent identity of two polynucleotide sequences, for example, can be performed by aligning the two sequences for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second nucleic acid sequences for optimal alignment and non-identical sequences can be disregarded for comparison purposes). In certain embodiments, the length of a sequence aligned for comparison purposes is at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or 100% of the length of the reference sequence. The nucleotides at corresponding nucleotide positions are then compared. When a position in the first sequence is occupied by the same nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which needs to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm. For example, the percent identity between two nucleotide sequences can be determined using methods such as those described in Computational Molecular Biology, Lesk, A. M., ed., Oxford University Press, New York, 1988; Biocomputing: Informatics and Genome Projects, Smith, D. W., ed., Academic Press, New York, 1993; Sequence Analysis in Molecular Biology, von Heinje, G., Academic Press, 1987; Computer Analysis of Sequence Data, Part I, Griffin, A. M., and Griffin, H. G., eds., Humana Press, New Jersey, 1994; and Sequence Analysis Primer, Gribskov, M. and Devereux, J., eds., M Stockton Press, New York, 1991; each of which is incorporated herein by reference. For example, the percent identity between two nucleotide sequences can be determined using the algorithm of Meyers and Miller (CABIOS, 1989, 4:11-17), which has been incorporated into the ALIGN program (version 2.0) using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. The percent identity between two nucleotide sequences can, alternatively, be determined using the GAP program in the GCG software package using an NWSgapdna.CMP matrix. Methods commonly employed to determine percent identity between sequences include, but are not limited to those disclosed in Carillo, H., and Lipman, D., SIAM J Applied Math., 48:1073 (1988); incorporated herein by reference. Techniques for determining identity are codified in publicly available computer programs. Exemplary computer software to determine homology between two sequences include, but are not limited to, GCG program package, Devereux, J., et al., Nucleic Acids Research, 12(1), 387 (1984)), BLASTP, BLASTN, and FASTA Altschul, S. F. et al., J. Molec. Biol., 215, 403 (1990)). [0369] Inhibit expression of a gene: As used herein, the phrase “inhibit expression of a gene” means to cause a reduction in the amount of an expression product of the gene. The expression product can be an RNA transcribed from the gene (e.g., an mRNA) or a polypeptide translated from an mRNA transcribed from the gene. Typically, a reduction in the level of an mRNA results in a reduction in the level of a polypeptide translated therefrom. The level of expression may be determined using standard techniques for measuring mRNA or protein. [0370] In vitro: As used herein, the term “in vitro” refers to events that occur in an artificial environment, e.g., in a test tube or reaction vessel, in cell culture, in a Petri dish, etc., rather than within an organism (e.g., animal, plant, or microbe). [0371] In vivo: As used herein, the term “in vivo” refers to events that occur within an organism (e.g., animal, plant, or microbe or cell or tissue thereof). [0372] Isolated: As used herein, the term “isolated” refers to a substance or entity that has been separated from at least some of the components with which it was associated (whether in nature or in an experimental setting). Isolated substances may have varying levels of purity in reference to the substances from which they have been associated. Isolated substances and/or entities may be separated from at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or more of the other components with which they were initially associated. In some embodiments, isolated agents are more than about 80%, about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more than about 99% pure. As used herein, a substance is “pure” if it is substantially free of other components. Substantially isolated: By “substantially isolated” is meant that the compound is substantially separated from the environment in which it was formed or detected. Partial separation can include, for example, a composition enriched in the compound of the present disclosure. Substantial separation can include compositions containing at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% by weight of the compound of the present disclosure, or salt thereof. Methods for isolating compounds and their salts are routine in the art. [0373] Label: The term “label” refers to a substance or a compound which is incorporated into an object so that the substance, compound or object may be detectable. [0374] Linker: As used herein, a linker refers to a group of atoms, e.g., 10-1,000 atoms, and can be comprised of the atoms or groups such as, but not limited to, carbon, amino, alkylamino, oxygen, sulfur, sulfoxide, sulfonyl, carbonyl, and imine. The linker can be attached to a modified nucleoside or nucleotide on the nucleobase or sugar moiety at a first end, and to a payload, e.g., a detectable or therapeutic agent, at a second end. The linker may be of sufficient length as to not interfere with incorporation into a nucleic acid sequence. The linker can be used for any useful purpose, such as to form conjugates, as well as to administer a payload, as described herein. [0375] Examples of chemical groups that can be incorporated into the linker include, but are not limited to, alkyl, alkenyl, alkynyl, amido, amino, ether, thioether, ester, alkylene, heteroalkylene, aryl, or heterocyclyl, each of which can be optionally substituted, as described herein. Examples of linkers include, but are not limited to, unsaturated alkanes, polyethylene glycols (e.g., ethylene or propylene glycol monomeric units, e.g., diethylene glycol, dipropylene glycol, triethylene glycol, tripropylene glycol, tetraethylene glycol, or tetraethylene glycol), and dextran polymers and derivatives thereof. Other examples include, but are not limited to, cleavable moieties within the linker, such as, for example, a disulfide bond (-S-S-) or an azo bond (-N=N-), which can be cleaved using a reducing agent or photolysis. Non-limiting examples of a selectively cleavable bond include an amido bond can be cleaved for example by the use of tris(2-carboxyethyl)phosphine (TCEP), or other reducing agents, and/or photolysis, as well as an ester bond can be cleaved for example by acidic or basic hydrolysis. [0376] Locked Nucleic Acids (LNAs): As used herein, “locked nucleic acids” or “LNAs” (also known as 2’-O,4’-C-methylene-bridged nucleic acid (2’,4’- BNA)) are artificial nucleic acid derivatives that contain a methylene bridge connecting the 2’-O with the 4’-C position in the furanose ring. This enables them to form a strictly N-type conformation that offers high binding affinity against complementary RNA. LNA also presents enzyme resistance, similar to other nucleic acid derivatives. LNAs are used for various gene silencing techniques, such as antisense, short interfering RNA, blocking of microRNA, and triplex-forming oligonucleotides. LNAs can be used, for example, in Splice Switching Oligonucleotides (SSOs) and LNA-based SSOs (LNA SSOs) are functional in vivo in mouse models. (See Shimo et. Al. Nucleic Acids Research, 42(12): 8174–8187 (2014), incorporated by reference herein in its entirety.) [0377] Modified: As used herein “modified” refers to a changed state or structure of a molecule of the disclosure. Molecules may be modified in many ways including chemically, structurally, and functionally. In some embodiments, the nucleic acid-based compositions of the present disclosure are modified by the introduction of non-natural nucleosides and/or nucleotides. [0378] Naturally occurring: As used herein, “naturally occurring” means existing in nature without artificial aid. [0379] Nucleic acid: The term "nucleic acid" as used herein, refers to a molecule comprised of one or more oligomeric polymers of at least 2 nucleoside monomers (nucleotide monomer when linked to another nucleoside. Each nucleoside monomer is further defined generally as comprising a sugar, nucleobase and backbone linker. Each of the sugar, nucleobase or backbone linker may be naturally occurring or synthetic. The nucleosides or nucleotides can be ribonucleotides, deoxyribonucleotides, or both. The term includes monomers and polymers of ribonucleotides and deoxyribonucleotides, with the ribonucleotides and/or deoxyribonucleotides being bound together, in the case of the polymers, via 5' to 3' linkages. However, linkages may include any of the linkages known in the art including, for example, nucleic acids comprising 5' to 3' linkages. The nucleotides may be naturally occurring or may be synthetically produced analogs that are capable of forming base-pair relationships with naturally occurring base pairs. Examples of non-naturally occurring bases that are capable of forming base-pairing relationships include, but are not limited to, aza and deaza pyrimidine analogs, aza and deaza purine analogs, and other heterocyclic base analogs, wherein one or more of the carbon and nitrogen atoms of the pyrimidine rings have been substituted by heteroatoms, e.g., oxygen, sulfur, selenium, phosphorus, and the like. [0380] Patient: As used herein, “patient” refers to a subject who may seek or be in need of treatment, requires treatment, is receiving treatment, will receive treatment, or a subject who is under care by a trained professional for a particular disease or condition. [0381] Peptide: As used herein, “peptide” is less than or equal to 50 amino acids long, e.g., about 5, 10, 15, 20, 25, 30, 35, 40, 45, or 50 amino acids long. [0382] Peptide nucleic acids (PNA): “Peptide nucleic acids” or “PNA” have a chemical structure similar to DNA or RNA but peptide bonds are used to link the nucleotides or nucleosides together. [0383] Pharmaceutically acceptable: The phrase “pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio. [0384] Pharmaceutically acceptable excipients: The phrase “pharmaceutically acceptable excipient,” as used herein, refers any ingredient other than the compounds described herein (for example, a vehicle capable of suspending or dissolving the active compound) and having the properties of being substantially nontoxic and non-inflammatory in a patient. Excipients may include, for example: antiadherents, antioxidants, binders, coatings, compression aids, disintegrants, dyes (colors), emollients, emulsifiers, fillers (diluents), film formers or coatings, flavors, fragrances, glidants (flow enhancers), lubricants, preservatives, printing inks, sorbents, suspensing or dispersing agents, sweeteners, and waters of hydration. Exemplary excipients include, but are not limited to: butylated hydroxytoluene (BHT), calcium carbonate, calcium phosphate (dibasic), calcium stearate, croscarmellose, crosslinked polyvinyl pyrrolidone, citric acid, crospovidone, cysteine, ethylcellulose, gelatin, hydroxypropyl cellulose, hydroxypropyl methylcellulose, lactose, magnesium stearate, maltitol, mannitol, methionine, methylcellulose, methyl paraben, microcrystalline cellulose, polyethylene glycol, polyvinyl pyrrolidone, povidone, pregelatinized starch, propyl paraben, retinyl palmitate, shellac, silicon dioxide, sodium carboxymethyl cellulose, sodium citrate, sodium starch glycolate, sorbitol, starch (corn), stearic acid, sucrose, talc, titanium dioxide, vitamin A, vitamin E, vitamin C, and xylitol. [0385] Pharmaceutically acceptable salts: The present disclosure also includes pharmaceutically acceptable salts of the compounds described herein. As used herein, “pharmaceutically acceptable salts” refers to derivatives of the disclosed compounds wherein the parent compound is modified by converting an existing acid or base moiety to its salt form (e.g., by reacting the free base group with a suitable organic acid). Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like. Representative acid addition salts include acetate, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptonate, hexanoate, hydrobromide, hydrochloride, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2- naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, toluenesulfonate, undecanoate, valerate salts, and the like. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like, as well as nontoxic ammonium, quaternary ammonium, and amine cations, including, but not limited to ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like. The pharmaceutically acceptable salts of the present disclosure include the conventional non-toxic salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. The pharmaceutically acceptable salts of the present disclosure can be synthesized from the parent compound which contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred. Lists of suitable salts are found in Remington’s Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418, Pharmaceutical Salts: Properties, Selection, and Use, P.H. Stahl and C.G. Wermuth (eds.), Wiley-VCH, 2008, and Berge et al., Journal of Pharmaceutical Science, 66, 1-19 (1977), each of which is incorporated herein by reference in its entirety. [0386] Pharmaceutically acceptable solvate: The term “pharmaceutically acceptable solvate,” as used herein, means a compound of the disclosure wherein molecules of a suitable solvent are incorporated in the crystal lattice. A suitable solvent is physiologically tolerable at the dosage administered. For example, solvates may be prepared by crystallization, recrystallization, or precipitation from a solution that includes organic solvents, water, or a mixture thereof. Examples of suitable solvents are ethanol, water (for example, mono-, di-, and tri-hydrates), N-methylpyrrolidinone (NMP), dimethyl sulfoxide (DMSO), N,N’- dimethylformamide (DMF), N,N’-dimethylacetamide (DMAC), 1,3-dimethyl-2- imidazolidinone (DMEU), 1,3-dimethyl-3,4,5,6-tetrahydro-2-(1H)-pyrimidinone (DMPU), acetonitrile (ACN), propylene glycol, ethyl acetate, benzyl alcohol, 2-pyrrolidone, benzyl benzoate, and the like. When water is the solvent, the solvate is referred to as a “hydrate.” [0387] Pharmacologic effect: As used herein, a “pharmacologic effect” is a measurable biologic phenomenon in an organism or system which occurs after the organism or system has been contacted with or exposed to an exogenous agent. Pharmacologic effects may result in therapeutically effective outcomes such as the treatment, improvement of one or more symptoms, diagnosis, prevention, and delay of onset of disease, disorder, condition or infection. Measurement of such biologic phenomena may be quantitative, qualitative or relative to another biologic phenomenon. Quantitative measurements may be statistically significant. Qualitative measurements may be by degree or kind and may be at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more different. They may be observable as present or absent, better or worse, greater or less. Exogenous agents, when referring to pharmacologic effects are those agents which are, in whole or in part, foreign to the organism or system. For example, modifications to a wild type biomolecule, whether structural or chemical, would produce an exogenous agent. Likewise, incorporation or combination of a wild type molecule into or with a compound, molecule or substance not found naturally in the organism or system would also produce an exogenous agent. [0388] The nucleic acid-based compositions of the present disclosure can comprise exogenous agents. Examples of pharmacologic effects include, but are not limited to, alteration in cell count such as an increase or decrease in neutrophils, reticulocytes, granulocytes, erythrocytes (red blood cells), megakaryocytes, platelets, monocytes, connective tissue macrophages, epidermal langerhans cells, osteoclasts, dendritic cells, microglial cells, neutrophils, eosinophils, basophils, mast cells, helper T cells, suppressor T cells, cytotoxic T cells, natural killer T cells, B cells, natural killer cells, or reticulocytes. Pharmacologic effects also include alterations in blood chemistry, pH, hemoglobin, hematocrit, changes in levels of enzymes such as, but not limited to, liver enzymes AST and ALT, changes in lipid profiles, electrolytes, metabolic markers, hormones or other marker or profile known to those of skill in the art. [0389] Physicochemical: As used herein, “physicochemical” means of or relating to a physical and/or chemical property. [0390] Preventing: As used herein, the term “preventing” refers to partially or completely delaying onset of an infection, disease, disorder and/or condition; partially or completely delaying onset of one or more symptoms, features, or clinical manifestations of a particular infection, disease, disorder, and/or condition; partially or completely delaying onset of one or more symptoms, features, or manifestations of a particular infection, disease, disorder, and/or condition; partially or completely delaying progression from an infection, a particular disease, disorder and/or condition; and/or decreasing the risk of developing pathology associated with the infection, the disease, disorder, and/or condition. [0391] Prognosing: As used herein, the term “prognosing” means a statement or claim that a particular biologic event will, or is very likely to, occur in the future. [0392] Progression: As used herein, the term “progression” or “cancer progression” means the advancement or worsening of or toward a disease or condition. [0393] Protein: A "protein" means a polymer of amino acid residues linked together by peptide bonds. The term, as used herein, refers to proteins, polypeptides, and peptides of any size, structure, or function. Typically, however, a protein will be at least 50 amino acids long. In some instances, the protein encoded is smaller than about 50 amino acids. In this case, the polypeptide is termed a peptide. If the protein is a short peptide, it will be at least about 10 amino acid residues long. A protein may be naturally occurring, recombinant, or synthetic, or any combination of these. A protein may also comprise a fragment of a naturally occurring protein or peptide. A protein may be a single molecule or may be a multi-molecular complex. The term protein may also apply to amino acid polymers in which one or more amino acid residues are an artificial chemical analogue of a corresponding naturally occurring amino acid. [0394] Protein expression: The term "protein expression" refers to the process by which a nucleic acid sequence undergoes translation such that detectable levels of the amino acid sequence or protein are expressed. [0395] Purified: As used herein, “purify,” “purified,” “purification” means to make substantially pure or clear from unwanted components, material defilement, admixture or imperfection. [0396] Sample: As used herein, the term “sample” or “biological sample” refers to a subset of its tissues, cells or component parts (e.g. body fluids, including but not limited to blood, mucus, lymphatic fluid, synovial fluid, cerebrospinal fluid, saliva, amniotic fluid, amniotic cord blood, urine, vaginal fluid and semen). A sample further may include a homogenate, lysate or extract prepared from a whole organism or a subset of its tissues, cells or component parts, or a fraction or portion thereof, including but not limited to, for example, plasma, serum, spinal fluid, lymph fluid, the external sections of the skin, respiratory, intestinal, and genitourinary tracts, tears, saliva, milk, blood cells, tumors, organs. A sample further refers to a medium, such as a nutrient broth or gel, which may contain cellular components, such as proteins or nucleic acid molecule. [0397] Signal Sequences: As used herein, the phrase “signal sequences” refers to a sequence which can direct the transport or localization of a protein. [0398] Single unit dose: As used herein, a “single unit dose” is a dose of any therapeutic administered in one dose/at one time/single route/single point of contact, i.e., single administration event. [0399] Similarity: As used herein, the term “similarity” refers to the overall relatedness between polymeric molecules, e.g. between polynucleotide molecules (e.g. DNA molecules and/or RNA molecules) and/or between polypeptide molecules. Calculation of percent similarity of polymeric molecules to one another can be performed in the same manner as a calculation of percent identity, except that calculation of percent similarity takes into account conservative substitutions as is understood in the art. [0400] Small interfering RNA (siRNA): As used herein “small interfering RNA” or “siRNA” mean a double-stranded RNA typically 20-25 nucleotides long involved in the RNA interference (RNAi) pathway and interfering with or inhibiting the expression of a specific gene. The gene is the target gene of the siRNA. A siRNA is usually about 21 nucleotides long, with 3' overhangs (e.g., 2 nucleotides) at each end of the two strands. A siRNA inhibits target gene expression by binding to and promoting the cleavage of one or more RNA transcripts of the target gene at specific sequences. Typically, in RNAi the RNA transcripts are mRNA, so cleavage of mRNA results in the down-regulation of gene expression. [0401] Splice-Switching Oligonucleotides (SSOs): As used herein, “Splice-switching oligonucleotides” or “SSOs” are oligonucleotides that modulate pre-mRNA splicing, can repair defective RNA, and restore the production of essential proteins. They can also generate novel proteins with desirable properties and regulate the presence of disease-related splice variant proteins. The latter outcome may be achieved by modulating alternative splicing of pre-mRNA. To modulate pre-mRNA splicing, SSOs may block RNA sequences that are essential for splicing and prevent the interaction of splicing factors, such as RNA- binding proteins, small nuclear RNAs and other components of the spliceosome, with the pre- mRNA. [0402] Split dose: As used herein, a “split dose” is the division of single unit dose or total daily dose into two or more doses. [0403] Stable: As used herein “stable” refers to a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and in some embodiments, capable of formulation into an efficacious therapeutic agent. [0404] Stabilized: As used herein, the term “stabilize”, “stabilized,” “stabilized region” means to make or become stable. [0405] Subject: As used herein, the term “subject” or “patient” refers to any organism to which a composition in accordance with the disclosure may be administered, e.g., for experimental, diagnostic, prophylactic, and/or therapeutic purposes. Typical subjects include animals (e.g., mammals such as mice, rats, rabbits, non-human primates, and humans) and/or plants. [0406] Substantially: As used herein, the term “substantially” refers to the qualitative condition of exhibiting total or near-total extent or degree of a characteristic or property of interest. One of ordinary skill in the biological arts will understand that biological and chemical phenomena rarely, if ever, go to completion and/or proceed to completeness or achieve or avoid an absolute result. The term “substantially” is therefore used herein to capture the potential lack of completeness inherent in many biological and chemical phenomena. [0407] Substantially equal: As used herein as it relates to time differences between doses, the term means plus/minus 2%. [0408] Substantially simultaneously: As used herein and as it relates to plurality of doses, the term means within 2 seconds. [0409] Suffering from: An individual who is “suffering from” a disease, disorder, and/or condition has been diagnosed with or displays one or more symptoms of a disease, disorder, and/or condition. [0410] Susceptible to: An individual who is “susceptible to” a disease, disorder, and/or condition has not been diagnosed with and/or may not exhibit symptoms of the disease, disorder, and/or condition but harbors a propensity to develop a disease or its symptoms. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition (for example, cancer) may be characterized by one or more of the following: (1) a genetic mutation associated with development of the disease, disorder, and/or condition; (2) a genetic polymorphism associated with development of the disease, disorder, and/or condition; (3) increased and/or decreased expression and/or activity of a protein and/or nucleic acid associated with the disease, disorder, and/or condition; (4) habits and/or lifestyles associated with development of the disease, disorder, and/or condition; (5) a family history of the disease, disorder, and/or condition; and (6) exposure to and/or infection with a microbe associated with development of the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition will develop the disease, disorder, and/or condition. In some embodiments, an individual who is susceptible to a disease, disorder, and/or condition will not develop the disease, disorder, and/or condition. [0411] Sustained release: As used herein, the term “sustained release” refers to a pharmaceutical composition or compound release profile that conforms to a release rate over a specific period of time. [0412] Synthetic: The term “synthetic” means produced, prepared, and/or manufactured by the hand of man. Synthesis of polynucleotides or polypeptides or other molecules of the present disclosure may be chemical or enzymatic. [0413] Targeted Cells: As used herein, “targeted cells” refers to any one or more cells of interest. The cells may be found in vitro, in vivo, in situ or in the tissue or organ of an organism. The organism may be an animal, in some embodiments, a mammal, or a human and most. In some embodiments, the targeted cells are cells of the kidney. The targeted cells can be a subpopulation of cells of the kidney. In some embodiments, the targeted cells are kidney tubule cells. [0414] Therapeutic Agent: The term “therapeutic agent” refers to any agent that, when administered to a subject, has a therapeutic, diagnostic, and/or prophylactic effect and/or elicits a desired biological and/or pharmacological effect. [0415] Therapeutically effective amount: As used herein, the term “therapeutically effective amount” means an amount of an agent to be delivered (e.g., nucleic acid, drug, therapeutic agent, diagnostic agent, prophylactic agent, etc.) that is sufficient, when administered to a subject suffering from or susceptible to an infection, disease, disorder, and/or condition, to treat, improve symptoms of, diagnose, prevent, and/or delay the onset of the infection, disease, disorder, and/or condition. [0416] Therapeutically effective outcome: As used herein, the term “therapeutically effective outcome” means an outcome that is sufficient in a subject suffering from or susceptible to an infection, disease, disorder, and/or condition, to treat, improve symptoms of, diagnose, prevent, and/or delay the onset of the infection, disease, disorder, and/or condition. [0417] Total daily dose: As used herein, a “total daily dose” is an amount given or prescribed in 24 hour period. It may be administered as a single unit dose. [0418] Treating: As used herein, the term “treating” refers to partially or completely alleviating, ameliorating, improving, relieving, delaying onset of, inhibiting progression of, reducing severity of, and/or reducing incidence of one or more symptoms or features of a particular infection, disease, disorder, and/or condition. For example, “treating” cancer may refer to inhibiting survival, growth, and/or spread of a tumor. Treatment may be administered to a subject who does not exhibit signs of a disease, disorder, and/or condition and/or to a subject who exhibits only early signs of a disease, disorder, and/or condition for the purpose of decreasing the risk of developing pathology associated with the disease, disorder, and/or condition. [0419] The phrase "a method of treating" or its equivalent, when applied to, for example, cancer refers to a procedure or course of action that is designed to reduce, eliminate or prevent the number of cancer cells in an individual, or to alleviate the symptoms of a cancer. "A method of treating" cancer or another proliferative disorder does not necessarily mean that the cancer cells or other disorder will, in fact, be completely eliminated, that the number of cells or disorder will, in fact, be reduced, or that the symptoms of a cancer or other disorder will, in fact, be alleviated. Often, a method of treating cancer will be performed even with a low likelihood of success, but which, given the medical history and estimated survival expectancy of an individual, is nevertheless deemed an overall beneficial course of action. [0420] Unmodified: As used herein, “unmodified” refers to any substance, compound or molecule prior to being changed in any way. Unmodified may, but does not always, refer to the wild type or native form of a biomolecule. Molecules may undergo a series of modifications whereby each modified molecule may serve as the “unmodified” starting molecule for a subsequent modification. [0421] Upregulation: The terms “upregulation” or “activation” as used in the context of a gene refer to an increase in the level of expression of the gene, an increase in the level of the polypeptide(s) encoded by the gene, the level bioactive gene product, or level of the RNA transcript(s) transcribed from the template strand of the gene as compared to the levels observed in the absence of the nucleic acid-based composition(s) described herein. The nucleic acid-based compositions of the present disclosure may have a direct upregulating effect on the expression of the target gene. The term “upregulation,” depending on the context, can also refer to the effect of a trans-acting activator of expression. VII. Equivalents and Scope [0422] Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments in accordance with the disclosure described herein. The scope of the present disclosure is not intended to be limited to the foregoing Description, but rather is as set forth in the appended claims. [0423] In the claims, articles such as “a,” “an,” and “the” mean one or more than one unless indicated to the contrary or otherwise evident from the context. Claims or descriptions that include “or” between one or more members of a group are considered satisfied if one, more than one, or all of the group members are present in, employed in, or otherwise relevant to a given product or process, unless indicated to the contrary or otherwise evident from the context. Throughout the Description and Claims, embodiments are provided in which exactly one member of the group is present in, employed in, or otherwise relevant to a given product or process. Throughout the Description and Claims, embodiments are provided in which more than one or all of the group members are present in, employed in, or otherwise relevant to a given product or process. [0424] Throughout the Description and Claims, use of the term “comprising” is intended to be open and contemplates or permits the inclusion of additional elements or steps. [0425] Where ranges are given, endpoints are included. Furthermore, it is to be understood that, unless otherwise indicated or otherwise evident from the context and understanding of one of ordinary skill in the art, values that are expressed as ranges can assume any specific value or subrange within the stated ranges in different embodiments of the disclosure, to the tenth of the unit of the lower limit of the range, unless the context clearly dictates otherwise. [0426] In addition, it is to be understood that any particular embodiment of the present disclosure that falls within the prior art may be explicitly excluded from any one or more of the claims. Since such embodiments are deemed to be known to one of ordinary skill in the art, they may be excluded even if the exclusion is not set forth explicitly herein. Any particular embodiment of the compositions of the disclosure (e.g., any nucleic acid; any method of production; any method of use; etc.) can be excluded from any one or more claims, for any reason, whether or not related to the existence of prior art. [0427] All cited sources, for example, references, publications, databases, database entries and art cited herein are incorporated into this application by reference even if not expressly stated in the citation. In case of conflicting statements of a cited source and the instant application, the statement in the instant application shall control. [0428] The disclosure is further illustrated by the following non-limiting examples. It is to be understood that the foregoing description and following examples are intended to illustrate and not limit the scope of the present disclosure, which is defined by the scope of the appended claims. EXAMPLES Example 1. Identification of GABPB1L Target Sequences [0429] The short nucleic acid sequences (tiles) were designed to represent the GABPB1L exon 9 coding sequence and splice sites. The coding sequence is represented by the genomic coordinates from the GRCh37/hg19 genomic assembly. The Hg19 coordinates for exon 9 of GABPB1L are chr15:50570829-50570981 (this sequence is provided as SEQ ID NO: 4). To include potential functional splice sites, the flanking DNA sequence was searched for splice site motifs using ESE finder 3.0 (website: krainer01.cshl.edu/cgi- bin/tools/ESE3/esefinder.cgi?process=home). Split site analysis identified chr15:50570778- 50570829 of the 3’ UTR and chr15:50570981-50570999 of the intron between exon 9 and exon 8 to be enriched for potential splicing motifs. Thus, the final target sequence used for oligonucleotide tiling design was chr15:50570778-50570999 (this sequence is provided as SEQ ID NO: 5). [0430] Figure 1 shows the exon-intron structure of GABPB1L and GABPB1S (not to scale). The inset shows exon 9 of GABPB1L as well as the flanking intron and UTR sequences. Putative splice motifs identified by computational analysis are represented by (*). The tiling window is defined by the vertical dotted lines, and candidate oligonucleotide sequences are represented by the horizontal black lines. Oligonucleotide designs that passed additional filtering criteria advanced to transfections in LN229-P2A-NLuc cells to test their ability to silence TERT expression. ASO Design [0431] Beginning at the start of SEQ ID NO: 5, every possible 15mer was generated with a sliding window of n=1, yielding 208 potential 15-mer oligonucleotide candidates. Those with a GC-content of <30% or >60% were filtered out. Remaining candidates were analyzed with GGRNA (website: GGRNA.dbcls.jp/) for target specificity. Oligos that aligned to 2 or more mRNA sequences were filtered out, leaving 100 oligo candidates. Each oligo candidate had Phosphorothioate backbones and Locked-Nucleic Acids (LNA) incorporated into their design to generate an antisense LNA oligo candidate (ASO) or a splice-switching LNA oligonucleotide candidate (SSO). The formula for each is designated below: ASO: +N*+N*+N*N*N*N*N*N*N*N*N*N*+N*+N*+N* (SEQ ID NO: 7) SSO: N*+N*N*+N*N*+N*N*+N*N*+N*N*+N*N*+N*N* (SEQ ID NO: 8) Where N = [A,C,G,T] DNA bases, * = a phosphorothioate backbone, and + = LNA. [0432] The hybridization temperature of each ASO and SSO to single stranded RNA was calculated with the exiqon online Tm tool (website: www.exiqon.com/ls/Pages/ ExiqonTMPredictionTool.aspx). Oligo candidates with Tm calculations of <58ºC and >91ºC removed. A positive control sequence, CTAACCAACAACGATC (SEQ ID NO: 9)) was used (Mancini et al, Cancer Cell 34(3):513-528 (2018), incorporated by reference herein in its entirety), as well as two scrambled negative control sequences were also included (TAO- 93/TSO-94 and TAO-94/TSO-95). The final list of ASOs (94) and SSO (95) candidates are listed in Tables 2 and 3. siRNA Design [0433] The target sequence for siRNA design included exon 9 of GABPB1L as well as the same portion of the 3’UTR that was scanned for ASOs (chr15:50570778-50570829). In addition, 11 base pairs of the flanking exon 8 sequence was included so that putative siRNAs spanning the exon 8-9 junction could be identified. The full cDNA sequence that was used to scan for siRNA is represented by SEQ ID NO: 6. [0434] This cDNA sequence was scanned for putative functional siRNA sequence using the selection criteria described in Reynolds et al., Nat. Biotechnol. 22(3):326-30 (2004) and Elbashir et al., Nature 411:494-498 (2001), each of which are incorporated herein in their entirety. Briefly, all sequence matches for Pattern 1 and Pattern 2 were identified. Pattern 1 was AA(N19)TT (SEQ ID NO: 10) and Pattern 2 was NA(N21) (SEQ ID NO: 11). N = any nucleotide. This yielded 81 siRNA candidate sequences. [0435] Candidate siRNA sequences with nucleotide repeats of four or greater, as well as with a GC content <30% or >60%, were filtered out. Additionally, BLAST searches were performed and each candidate with a sequence match of >18 nucleotides or matching more than 5 genes (E-score <1) were also removed. This resulted in a final candidate list of 39 duplex candidates shown in Table 9 above. Dharmacon (website: dharmacon.horizondiscovery.com) siGenome SMART pools were ordered as control siRNAs. siGABPB1 (M-013083-01-0005), siTERT (M-003547-02-0005), and non-targeting siCTRL (D-001206-13-05), were used as positive and negative controls respectively. Example 2. Nucleic Acid Screening Synthesis [0436] ASOs, SSOs, and ASO duplexes (in this instance, siRNAs) were all ordered from Integrated DNA Technologies (website: www.idtdna.com). ASOs and SSOs were ordered at 100nmol scale with standard desalting. siRNA sense and antisense RNA strands were ordered at 100nmol scale and underwent hybridization and concentration normalization prior to delivery. siRNA Primary Screen [0437] All candidate siRNAs were initially transfected into a glioblastoma (GBM) reporter cell line (LN229-P2A-NLuc) expressing NanoLuc Luciferase endogenously through a P2A fusion with the endogenous TERT gene. This knock-in reporter is on the same allele as the C228T TERTp mutation and provides a readout of TERT mRNA expression changes via luciferase activity. LN229-P2A-NLuc was prepared as follows: [0438] The NanoLuc Luciferase protein was generated as an in-frame fusion with the endogenous TERT gene on the same allele containing the C228T TERT promoter mutation. A homologous recombination knock-in approach was used. The final 300bp of the TERT coding sequence, followed by a P2A self-cleaving peptide sequence (derived from Porcine teschovirus-12A), followed by NanoLuc Luciferase, followed by the first 300bp of the TERT 3’ UTR were cloned into the pUC57 vector in sequence (plasmid referred to as TERT- P2A-NLuc-TERT). [0439] In addition, CRISPR guide RNAs designed to cut at the TERT coding sequence- 3’UTR junction were cloned into the px459 vector (Genscript, Piscataway, NJ, Cat No. SC1814 ) and their sequence was confirmed by Sanger sequencing (plasmid referred to as TERT-g1-px459). [0440] To generate the stable knock-in cell line, 1 million LN229 cells were seeded in T25 flasks 24 hours prior to transfection. TERT-g1-px459 and TERT-P2A-NLuc-TERT plasmids were co-transfected using the lipofectamine 3000 system (ThermoFisher, Waltham, MA, Cat No. L3000001). 48 hours post transfection, 1 μg/ml of Puromycin was added to the cells to select for cells that had successfully taken up the TERT-g1-px459 plasmid. [0441] The cells were propagated for 6 days in the presence of puromycin. On day 6, the cells were trypsinized. Half of the cells were re-seeded in 96 well plates at a density of 20 cells/well while the other half were used to collect genomic DNA. Junction PCR was performed on the genomic DNA to detect the presence of cells containing the correct knock- in at the TERT locus. After two weeks of expansion, NanoLuc Luciferase was measured in each well of the 96-well plate and the wells with a positive signal were advanced to single cell cloning. [0442] Single-cell clones containing the correct integration were derived and frozen down for future use (termed LN229-P2A-Nluc hereafter). These cells had the P2A-NanoLuc Luciferase fusion knock-in confirmed by Junction-PCR. In addition, siRNA of TERT confirmed a reduction in NLuc signal as would be expected from a successful knock-in. [0443] Cells were seeded at 4,000 cells/well in 96-well plates 24 hours prior to transfection. siRNAs were transfected at 50nM. All transfections were performed with Dharmafect1 (Dharmacon: T-2001-01), with 0.15uL of Dharmafect 1 being used for each well. [0444] NanoLuc Luciferase was measured 48 hours and 72 hours post transfection using the Live-Cell Nano Glo kit (Promega N2011). This analysis was performed three independent times, the data was standardized to a zero-centered gaussian distribution and z-scores were combined for replicate wells to calculate mean and standard deviation effect sizes. ASO Primary Screen [0445] All candidate ASOs were initially transfected into the LN229 GBM cell line, containing the C228T TERTp mutation. Cells were seeded at 4,000 cells/ well in 96-well plates 24 hours prior to transfection. The following day, ASOs were transfected at 25nM. All transfections were performed with Dharmafect1 (Dharmacon, Lafayette, CO, T-2001-01), with 0.15uL of Dharmafect 1 being used for each well. 72 hours post transfection, the cells were harvested for RNA and changes in TERT mRNA were determined by RT-qPCR analysis (GUSB primers were used as an internal control). [0446] Figure 2 provides a set of charts showing the expression fold change, nano luciferase and fluorescence (590 nm) for the samples at 72 hours. LN229 GBM cells (TERTp mutant) and LN229-P2A-NLuc cells were transfected with 50nM of candidate siRNA. 72 hours post drug treatment, cell lysates were harvested and TERT mRNA was measured via RT-qPCR (ThermoFisher, Waltham, MA, Cat. No. A25600). Replicate plates were used to measure cell viability (ThermoFisher PrestoBlue: A13261), and the LN229- P2A-NLuc cell line was used to measure NanoLuc Luciferase. All treatment samples were normalized to the vehicle control of their respective cell type. Error bars represent a standard deviation of three replicates. [0447] TSiR-1, TSiR-37, TSiR-26, and TSiR-34 were selected as hits for displaying a reduction in NanoLuc Luciferase activity at either 48 or 72 hours. In addition, TSiR-6 was randomly selected from the candidates that had no effect on reporter activity. These 5 siRNAs advanced to hit confirmation analysis. siRNA hit Confirmation [0448] 24-hours prior to transfection, 4,000 cells/well of LN229 GBM cells were seeded into two 96-well plates. A matching plate of LN229-P2A-NLuc cells were also seeded. Triplicate wells were used for each condition, and each siRNA was transfected as described above. 72-hours post transfection, RNA was harvested from plate 1, plate 2 was used to measure cell-viability (ThermoFisher PrestoBlue: A13261), and the LN229-P2A-NLuc plate was used to measure NanoLuc Luciferase. [0449] RT-qPCR analysis was performed on all collected RNA lysates to determine relative expression changes in GABPB1L and TERT mRNA (GUSB primers were used as an internal control). siRNA Cell Panel Analysis [0450] To test if any of the siRNA hits could reduce TERT expression selectively in cancer cells harboring TERTp mutations, we performed additional transfections in a panel of two TERTp-mutant and two TERTp wild-type cell lines. LN18 is a GBM cell line that expresses TERT from the wild-type promoter, and 293T cells are TERTp wild-type cells derived from embryonic kidney cells. LN229 is GBM cell line and HepG2 is a Hepatocellular carcinoma cell line, both of which are positive for the C228T TERTp mutation. [0451] Each cell line underwent a transfection optimization to determine the ideal amount of Dharmafect 1 transfection reagent to achieve maximal GABPB1L knockdown while minimizing cell toxicity. It was found that 0.2μL/well was ideal for LN229 and LN18, 0.3μL/well was optimal for 293T, and 0.4μL/well was optimal for HepG2 cells. [0452] TSiR-1, TSiR-37, TSiR-26, and TSiR-34 were transfected at 50nM into the four cell lines in 96-well plates. siCTRL (scrambled non-targeting) and siGABPB1 were used as negative and positive control siRNAs respectively. 72 hours post transfection, the cells were harvested for RNA and changes in GABPB1L and TERT mRNA (Figure 3 and Figure 4, respectively) were determined by RT-qPCR analysis. Both TSiR-1 and TSiR-34 achieved a greater TERT reduction in the two TERTp mutant lines when compared to the TERTp wild- type cell lines. For TSiR-1, this effect was particularly pronounced as the knockdown efficiency of GABPB1L was greatest in LN18 (WT) and weakest in HepG2 (Mut). ASO Cell Panel Analysis [0453] TAO-21, TAO-22, TAO-37, and TAO-40 were selected from the primary screen as hits for displaying greater than 60% reduction in TERT mRNA at 72 hours. To test if any of the ASO hits could reduce TERT expression selectively in cancer cells harboring TERTp mutations, we performed additional transfections in a panel of two TERTp-mutant and two TERTp wild-type cell lines. LN18 is a GBM cell line that expresses TERT from the wild- type promoter, and 293T cells are TERTp wild-type cells derived from embryonic kidney cells. LN229 is GBM cell line and HepG2 is a Hepatocellular carcinoma cell line, both of which are positive for the C228T TERTp mutation. [0454] The four ASO hits were transfected at 25nM into the four cell lines in 96-well plates. A scrambled non-targeting LNA-ASO (Scr) was used as a negative control. 72 hours post transfection, the cells were harvested for RNA and changes in TERT mRNA were determined by RT-qPCR analysis (Figure 5). TAO-21, TAO-22, and TAO-37 achieved a greater TERT reduction in the two TERTp mutant lines when compared to the TERTp wild- type cell lines. Example 3. Identification of ASOs [0455] All possible 20-mer GABPB1-hybridizing oligonucleotides (MOE, 5-10-5) were designed to target GABPB1L or total GABPB1 (both the GABPB1L and GABPB1S isoforms). The GABPB1-hybridizing oligonucleotides that were designed to target GABPB1L are shown in Table 5 and the GABPB1-hybridizing oligonucleotides that were designed to target total GABPB1 are shown in Table 6. Screen of 150 ASOs [0456] Based on the bioinformatics analysis of the oligonucleotides in Table 5 and Table 6, 150 ASOs (Table 11) were selected for screening. The candidate ASOs were transfected into Hep3B cells which were seeded at 15,000 cells/well in 96-well plates. The following day, ASOs were transfected at 20 nM or 2 nM. 24 hours post transfection, the cells were harvested and GABPB1L was measured with a bDNA assay. The mean relative GABPB1L mRNA level (“Relative mRNA Expression”) for the 20 nM and 2 nM doses for each ASO tested is shown in Table 11 as well as the cell viability for the 20 nM dose. Table 11. Mean Relative GABPB1L mRNA
Figure imgf000253_0001
Figure imgf000254_0001
Figure imgf000255_0001
Figure imgf000256_0001
Figure imgf000257_0001
[0457] Based on the results above, 50 ASOs were selected for additional studies. Screen of 50 ASOs [0458] 50 ASOs were selected from the screen of the 150 ASOs. The candidate ASOs were transfected into Hep3B cells which were seeded at 15,000 cells/well in 96-well plates. The following day, ASOs were transfected at 20 nM or 5 nM. 72 hours post transfection, the cells were harvested and GABPB1L and TERT mRNA was measured by RT-qPCR. The knockdown percentage for GABPB1L and TERT are in shown in Table 12 as well as the cell viability for the 5 nM dose. Table 12. GABPB1L and TERT mRNA Expression
Figure imgf000257_0002
Figure imgf000258_0001
Microwalk Redesign and Screen [0459] TAO-A2641.1, TAO-A4902.1, TAO-A5551.1, TAO-A5552.1, TAO-A6451.1, TAO-A6453.1, TAO-A6529.1, TAO-A6577.1, TAO-A7186.2, TAO-A7210.1, TAO- A7215.1, TAO-A7441.1, TAO-A7797.1, TAO-A8210.1, TAO-A10683.1, TAO-A11082.1, and TAO-A11083.1 (collectively referred to as “17 ASOs”) were selected from the screen of the 50 ASOs described in Table 12 for further screening and to design the additional 16-mer, 18-mer and 20-mer ASOs shown in Table 8. [0460] The 17 ASOs and the 63 ASOs from Table 12, (a total of 80 ASOs) were transfected into Hep3B cells which were seeded at 15,000 cells/well in 96-well plates. The following day, ASOs were transfected at 20 nM or 5 nM. 24 hours post transfection, the cells were harvested and mRNA levels were measured by bDNA assay for hsGABPB1L, hsCDKN1A (hsGABPB1L silencing should lead to an upregulation of hsCDKN1A) and hsGAPDH. The mean value (MV) of the remaining mRNA levels GAPDH percentage (GAPDH %) for each ASO is shown in Table 13. Table 13. GABPB1L, hsCDKN1A Expression and GAPDH Percentage
Figure imgf000259_0001
Figure imgf000260_0001
Figure imgf000261_0001
Example 4. In Vitro Transfection Studies [0461] A series of screens were performed to identify oligonucleotides that decrease GABPB1L (the beta1L tetramer-forming isoform of GABP) and/or TERT mRNA expression in vitro. TAO-95.1 to TAO-122.1: U87, Huh7, LN229 and 293T Cells [0462] TAO-95.1 to TAO-122.1 were synthesized and transfected into U87 and Huh7 cells that are both positive for the C228T mutation. An initial screen using a PrestoBlue® assay (ThermoFisher) was performed to determine cell viability and qPCR was used to measure the levels of GABPB1L and TERT mRNA. The average cell viability and the average TERT mRNA fold change after 72 hours is provided in Table 14. A scrambled non- targeting LNA-ASO (Scr) was used as a negative control and TAO-37.1 was used as a positive control. Table 14. Cell Viability and TERT mRNA Fold Change in Huh7 and U87 Cells
Figure imgf000261_0002
Figure imgf000262_0001
[0463] Based on the results from the U87 and Huh7 cell line study, TAO-95.1, TAO-97.1, TAO-104.1, TAO-106.1, TAO-115.1, TAO-116.1, TAO-118.1 and TAO-120.1 were selected for additional studies in LN229 (GBM cell line) and 293T (TERTp wild-type cells derived from embryonic kidney cells) cells. The average cell viability and the average TERT mRNA fold change for the LN229 and the 293T cells after 72 hours is provided in Table 15. A comparison of the ASOs across the 4 cell lines (Huh7, U87, LN229 and 293T) is shown in Table 16. A scrambled non-targeting LNA-ASO (Scr) was used as a negative control and TAO-37.1 was used as a positive control. Table 15. Cell Viability and TERT mRNA Fold Change in LN229 and 293T Cells
Figure imgf000262_0002
Table 16. Cell Viability and TERT mRNA Fold Change Across Huh7, U87, LN229 and 293T Lines
Figure imgf000263_0001
TAO-123.1 to TAO-139.1: Huh7 and LN229 Cells [0464] TAO-123.1 to TAO-139.1 where were synthesized and transfected into Huh7 and LN229 cells that are both positive for the C228T mutation. TAO-123.1 and TAO-125.1 are locked nucleic acids (LNAs) that have a backbone that includes phosphorothioate (Ps) and phosphodiester (Po) bonds, TAO-124.1 is an LNA that has a backbone that includes Ps bonds, and TAO-126.1 to TAO-139.1 include 2’-O-(2-Methoxyethyl)-modified nucleotides (2-MOE) and backbones that include Ps and Po bonds. An initial screen using a PrestoBlue® assay (ThermoFisher) was performed to determine cell viability and qPCR was used to measure the levels of GABPB1L and TERT mRNA. The average cell viability, and the average GABPB1L mRNA and TERT mRNA fold change at 48 hours is provided in Table 17. A scrambled non-targeting LNA-ASO (ScrASO) was used as a negative control, and TAO-115.1 and TAO-95.1 were used as a positive control. Table 17. Cell Viability, TERT mRNA and GABPB1L Fold Change in Huh7 and LN229 Cells (48 Hours)
Figure imgf000263_0002
Figure imgf000264_0001
[0465] Based on the results from the 48 hour results in the LN229 and Huh7 study, TAO- 123.1, TAO-125.1, TAO-130.1, TAO-131.1, TAO-132.1, TAO-135.1, TAO-137.1 and TAO- 139.1 were selected for a follow-up time course study. The average cell viability, and the average GABPB1L mRNA and TERT mRNA fold change at 96 hours is provided in Table 18. Provided in Table 19 is a summary of the time course data for the LN229 cells. A scrambled non-targeting LNA-ASO (ScrASO) was used as a negative control along with an untreated sample, and TAO-115.1 and TAO-95.1 were used as a positive control. [0466] Cell toxicity was found to be associated with highly variable TERT expression and the LNA ASOs (TAO-123.1 to TAO-126.1) were found to have lower cell viability as compared to the non-LNA ASOs. Knockdown of GABPB1 expression over the time course study depended on the ASO, but TAO-137.1, TAO-139.1 and TAO-132.1 were found to have the best stable effects on GABPB1 and TERT expression over the length of the study. Table 18. Cell Viability, TERT mRNA and GABPB1L Fold Change in Huh7 and LN229 Cells (96 Hours)
Figure imgf000264_0002
Figure imgf000265_0001
Table 19. Time Course Summary in LN229 Cells
Figure imgf000265_0002
Example 5. Gymnotic Dose-Response Study [0467] Three ASOs (TAO-115.1, TAO-37.1 and TAO-98.1) were selected for gymnotic dose-response studies with 9-point dose response curves (80 uM to 0.313 uM) in U87, U251 and LN229 cells that are positive for the C228T mutation and LN18 cells (LN18 is a GBM cell line that expresses TERT from the wild-type promoter). A PrestoBlue® assay (ThermoFisher) was performed to determine cell viability and qPCR was used to measure the levels of GABPB1L and TERT mRNA at 96 hours. Table 20 shows the average cell viability where DMSO was used as the vehicle only control and the results are shown as percent viability from vehicle. Shown in Table 21 are the average GABPB1L mRNA fold change as compared to the vehicle only and Table 22 are the average TERT mRNA fold change as compared to the vehicle only. For TAO-37.1 and TAO-115.1, the mRNA expression fold change for both GABPB1L and TERT was lowest for the higher dose and was highest at the lowest dose. For TAO-98.1 this trend was only seen for TERT expression. Table 20. Cell Viability for TAO-37.1, TAO-98.1, TAO-115.1
Figure imgf000265_0003
Figure imgf000266_0002
Figure imgf000266_0001
Figure imgf000266_0003
Figure imgf000267_0001
Table 22. TERT mRNA Fold Change for TAO-37.1, TAO-98.1, TAO-115.1
Figure imgf000267_0002
Figure imgf000268_0001
Example 6. In Vivo Study [0468] Various in vivo studies were conducted to evaluate a subset of the ASOs described herein. CED Delivery of ASO [0469] Three ASOs (TAO-115.1, TAO-37.1 and TAO-98.1) were selected for in vivo studies in mice. Mice were inoculated on day 1 with tumor and then administered 5 uL of TAO-115.1, TAO-37.1 and TAO-98.1 or a control of scrambled non-targeting LNA-ASO (ScrASO) by convection enhanced delivery (CED). On day 6 the tumors were harvested and analyzed by RT-qPCR. The average and median relative expression (Fold Change to ScrASO) of GABPB1 and TERT are shown in Table 23 and Table 24 respectfully. Table 23. GABPB1 mRNA Fold Change for TAO-37.1, TAO-98.1, TAO-115.1
Figure imgf000268_0002
Table 24. TERT mRNA Fold Change for TAO-37.1, TAO-98.1, TAO-115.1
Figure imgf000269_0001
ITu Delivery of ASO [0470] TAO-115.1 was selected for in vivo studies in mice comparing a single dose or multi-dosing regimen. For the single dose regimen, mice were inoculated with tumor, BLI was measured at day 7 and day 12, and then on day 12 the mice were administered by intratumoral (ITu) delivery a single dose of 300 ug ScrASO, 300 ug of TAO-115 or 500 ug of TAO-115. For the multi-dosing regimen, mice were inoculated with tumor, BLI was measured at day 7 and day 12, and then on day 12, day 14 and day 16 the mice were administered by intratumoral (ITu) delivery a dose of 300 ug ScrASO, 200 ug of TAO-115.1 or 300 ug of TAO-115.1. For both the single and multi-dose studies at day 19 the bioluminescence (BLI) was measured and tumors were harvested and analyzed by RT-qPCR. Dose-dependent reduction of GABPB1L was observed and the results are shown in Table 25. Shown Table 25 are the fold change for GABPB1 mRNA and TERT mRNA as compared to ScrASO. Table 25. TERT and GABPB1 mRNA Fold Change
Figure imgf000269_0002
Example 7. Additional In Vitro Studies [0471] 16 ASOs were tested for their effects on cell viability, GABPB1L and TERT mRNA expressions in LN229 (TERTp mutant) and Huh7 (TERTp mutant) cells. Cells were seeded at 4,000 cells/well in 96-well plates 24 hours prior to transfection. [0472] The following day, ASOs were transfected at 25nM. All transfections were performed with Dharmafect1 (Dharmacon, Lafayette, CO, T-2001-01), with 0.15uL of Dharmafect 1 being used for each well. 72 hours post transfection, the cells were assayed for viability using the PrestoBlue assay (Thermofisher) and then harvested for RNA (ThermoFisher, Waltham, MA, Cat. No. A25600). Changes in GABPB1L and TERT mRNA expression were determined by RT-qPCR analysis (GUSB primers were used as an internal control). The mean value (MV) of cell viability, GABPB1L mRNA, and TERT mRNA data are shown in Table 26. Table 26. Cell Viability and GABPB1 and TERT mRNA % Change
Figure imgf000270_0001
* The negative value means the mRNA level was reduced and the positive value means the mRNA level was increased.

Claims

CLAIMS 1. An oligonucleotide capable of hybridizing to a target sequence of a GABPB1 gene (DNA) or gene product (RNA), wherein said target sequence comprises a sequence located in exon 9, the intron between exon 8 and exon 9, or the exon 8 and exon junction of a GABPB1 or the target sequence comprises a sequence targeting both GABPB1S and GABPB1L isoforms of GABPB1, and wherein said target sequence is any of SEQ ID NOs: 308-346 and 12424-24399.
2. The oligonucleotide of claim 1, comprising a sequence of 8-20 nucleotides in length and wherein hybridization is over at least 50% of the oligonucleotide sequence.
3. The oligonucleotide of claim 2, comprising a sequence of 8-20 nucleotides in length and wherein hybridization is over at least 90% of the oligonucleotide sequence.
4. The oligonucleotide of claim 3, comprising a sequence of 8-20 nucleotides in length and wherein hybridization is over at least 99% of the oligonucleotide sequence.
5. The oligonucleotide of any of claims 1-4, comprising one or more modified nucleobases, sugars or linkers.
6. The oligonucleotide of any one of claims 1-5, which is a 15-mer, a 16-mer, 18-mer or a 20-mer.
7. The oligonucleotide of any one of claims 1-6, where the oligonucleotide comprises any of SEQ ID NOs: 112-205, 300-307, 475-12189, 12340-12360, and 24440-24444.
8. The oligonucleotide of any one of claims 1-6, where the oligonucleotide comprises any of SEQ ID NOs: 6019, 11151, 11547, 11550, 11553, 11554, 12154, 12340, 12343, 12346, 12347, 12348, 12353, 12357, 12358, and 12359.
9. The oligonucleotide of any one of claims 1-6, where the oligonucleotide comprises any of SEQ ID NOs: 131, 133, 148, 151, 24403, and 24420.
10. The oligonucleotide of any one of claims 1-9, wherein oligonucleotide is an antisense oligonucleotide (ASO) that is a fully modified 3-9-3 LNA compound.
11. The oligonucleotide of any one of claims 1-9, comprising one or more phosphorothioate (Ps) linkages.
12. The oligonucleotide of any one of claims 1-9, comprising one or more phosphodiester (Po) linkages.
13. The oligonucleotide of any one of claims 1-9, wherein one or more cytosine bases is modified to be 5-methylcytosine.
14. The oligonucleotide of any one of claims 1-9, comprising 5’-terminal and 3’-terminal 2’MOE modified nucleotides.
15. The oligonucleotide of claim 14, further comprising at least one internal 2’MOE modified nucleotide.
16. An antisense oligonucleotide (ASO) comprising the sequence of any one of SEQ ID NO: 19-111, 430-474, 12190-12339, and 12361-12423.
17. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12335.
18. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12338.
19. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12363.
20. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12372.
21. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12376.
22. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12379.
23. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12382.
24. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12383.
25. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12384.
26. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12389.
27. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12393.
28. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12394.
29. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12395.
30. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12413.
31 The ASO of claim 16, where the ASO comprises SEQ ID NO: 12421.
32. The ASO of claim 16, where the ASO comprises SEQ ID NO: 12422.
33. A double-stranded small inhibitory RNA (siRNA) comprising the sequence of any one of SEQ ID NOs: 386-424.
34. The siRNA of claim 33, wherein said sequence comprises SEQ ID NO: 386.
35. The siRNA of claim 33, wherein said sequence comprises SEQ ID NO: 411.
36. The siRNA of claim 33, wherein said sequence comprises SEQ ID NO: 419.
37. The siRNA of claim 33, wherein said sequence comprises SEQ ID NO: 422.
38. A pharmaceutical composition comprising an oligonucleotide of any one of claims 1- 15, an ASO of any one of claims 16-32, or a siRNA of any one of claims 33-37, and a pharmaceutically acceptable carrier or diluent.
39. A method of reducing the activity of GA binding protein transcription factor beta 1 (GABPB1) in a cell, comprising exposing said cell to the pharmaceutical composition of claim 38
40. A method of reducing the expression of telomerase reverse transcriptase (TERT) in a cell having a mutant TERT promoter with one or more somatic mutations, comprising exposing said cell to the pharmaceutical composition of claim 38.
41. A method of decreasing the level of one or more GABPB1 gene product(s) expressed in a cell, tissue, organ or organism comprising contacting said cell, tissue, organ or organism with a pharmaceutical composition of claim 38.
42. The method of claim 41, wherein the GABPB1 gene product decreased is total GABPB1.
43. A method of decreasing the level of one or more TERT gene product(s) expressed in a cell, tissue, organ or organism comprising contacting said cell, tissue, organ or organism with a pharmaceutical composition of claim 38.
44. A method of treating cancer in a subject comprising administering to the subject a pharmaceutical composition of claim 38.
45. A use of the oligonucleotide of any one of claims 1-15, an ASO of any one of claims 16-32, or a siRNA of any one of claims 33-37 for the manufacture of a pharmaceutical composition for treating cancer.
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