EP4669750A2

EP4669750A2 - HIBITORS OF SYNAPTOGYRIN-3 EXPRESSION

Info

Publication number: EP4669750A2
Application number: EP24706719.2A
Authority: EP
Inventors: Ana Rita SANTOS; Patrik Verstreken
Original assignee: Jay Therapeutics; Katholieke Universiteit Leuven; Vlaams Instituut voor Biotechnologie VIB
Current assignee: Jay Therapeutics; Katholieke Universiteit Leuven; Vlaams Instituut voor Biotechnologie VIB
Priority date: 2023-02-21
Filing date: 2024-02-20
Publication date: 2025-12-31
Also published as: WO2024175586A3; WO2024175586A2

Abstract

The invention relates to identification of regions within the synaptogyrin-3 RNA sequence that are targetable by oligonucleotide inhibitors such as siRNA molecules. The synaptogyrin-3 inhibitors disclosed herein are provided for use as a medicament in general, and for treating or inhibiting progression of tauopathies or symptoms of tauopathies in particular.

Description

INHIBITORS OF SYNAPTOGYRIN-3 EXPRESSION

FIELD OF THE INVENTION

The invention relates to regions within the synaptogyrin-3 RNA sequence that are targetable by oligonucleotide inhibitors such as siRNA molecules. The synaptogyrin-3 inhibitors disclosed herein are provided for use as a medicament in general, and for treating or inhibiting progression of tauopathies or symptoms of tauopathies in particular.

BACKGROUND TO THE INVENTION

Tau pathology is associated with more than twenty neurodegenerative diseases, including Alzheimer's disease (Wang & Mandelkow 2016 Nat Rev Neurosci 17:5-21). Hyperphosphorylation or mutation of the microtubule-associated protein Tau is common to all of these diseases, collectively termed Tauopathies, and filamentous inclusions of hyperphosphorylated Tau are hallmark pathologies of Alzheimer's disease and other Tauopathies (Ballatore et al 2007 Nature Reviews Neuroscience 8:663-672). Tau pathology is not merely a byproduct of other pathological pathways, but is a key mediator of neurotoxicity itself (Roberson et al 2007 Science 316:750-754; Hutton et al 1998 Nature 393:702-705; Caffrey & Wade- Martins 2007 Neurobiol Dis 27:1-10; Le Guennec et al 2016 Molecular Psychiatry 1-7). Under physiological conditions, Tau is expressed in neurons and is bound to axonal microtubules. However, under pathological conditions, mutations in Tau (for example, in Frontotemporal dementia with parkinsonism(FTDP)-17) or abnormal phosphorylation of Tau (for example in sporadic Alzheimer's disease) decrease its microtubule binding affinity (Hong et al 1998 Science 282:1914-1917; Wang & Mandelkow 2016 Nat Rev Neurosci 17:5-21), leading to its dissociation from axonal microtubules and subsequent mislocalization to synapses (Spires-Jones & Hyman 2014 Neuron 82:756-771; Tai et al 2012 Am J Pathol 181:1426-1435; Tai et al 2014 Acta Neuropathol Commun 2:146). This mislocalisation of soluble Tau plays a key role in perturbing synaptic function in early disease stages, which may contribute to subsequent synapse loss and neurodegeneration.

In addition to the reported post-synaptic localization of pathological Tau (Hoover et al 2010 Neuron 68:1067-1081; Ittner et al 2010 Cell 142:387-397; Zhao et al 2016 Nat Med 22:1268-1276), it was previously shown that hyperphosphorylated Tau species accumulate on pre-synaptic vesicles isolated from Alzheimer's disease patient brain. This suggests that the pre-synaptic pathway also contributes to synaptic dysfunction in human neurodegenerative diseases associated with Tau. Using unbiased proteomic and genetic approaches, it was found that the transmembrane synaptic vesicle protein Synaptogyrin-3 mediates the association of Tau with synaptic vesicles in vitro and in vivo (W02019/016123). Reduction of Drosophila Synaptogyrin or murine Synaptogyrin-3 levels in neurons from fly and mouse models of tauopathy reduced the association of Tau with synaptic vesicles, and subsequently rescued Tau-induced defects in vesicle mobility and neurotransmitter release. These findings identified Synaptogyrin-3 as a novel Tau interactor that mediates synaptic dysfunction associated with Tau (W02019/016123). It would thus be advantageous to develop specific Synaptogyrin- 3 inhibitors for use in treating various Tauopathies including Alzheimer's disease, conditions for which there continues to be a high unmet need for therapies. Impacting Tau-mediated pathways at the presynapse is highly relevant given that neurodegeneration is thought to begin with loss of presynaptic terminals and proceed retrograde in a dyeing-back process (Yoshiyama et al 2007 Neuron 53:337-351).

SUMMARY OF THE INVENTION

The inventors of current application have found that some subsequences within the Synaptogyrin-3 mRNA transcript are significantly more accessible for oligonucleotides such as RNAi molecules and therefore are preferred target regions for designing oligonucleotides suitable for or capable of reducing the expression and/or activity of Synaptogyrin-3. The borders of those identified target regions were determined by transcript-walking.

Therefore the application provides an oligonucleotide of 10 to 70 nucleotides in length, comprising a contiguous nucleotide sequence of at least 10 contiguous nucleotides in length, the contiguous nucleotide sequence being at least 90% complementary to an equal length portion of a target region within the Synaptogyrin-3 transcript as depicted in SEQ ID No. 1, wherein the target region is comprised between nucleobase 205 and 265, 255 and 348, 338 and 387, 369 and 433, 422 and 531, 603 and 656, 641 and 714, 717 and 768, 1150 and 1600, 1743 and 1868 or between nucleobase 1865 and 2026 of SEQ. ID No. 1 and wherein the endpoints are included. In one embodiment, the oligonucleotide can bind to the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. In another embodiment, said binding of the oligonucleotide to said synaptogyrin-3 mRNA transcript can reduce the expression and/or activity of synaptogyrin-3.

In one embodiment, the oligonucleotide is a double stranded nucleic acid molecule, more particularly an RNAi molecule or an RNA duplex, even more particularly the RNAi molecule is an siRNA, a divalent siRNA or a shRNA. In another embodiment, the oligonucleotide is a single stranded nucleic acid molecule, more particularly the antisense portion of an RNAi molecule.

In another embodiment, the sense and/or antisense strand of the oligonucleotide of the application comprises between 15 and 25 nucleotides in length, more particularly the antisense strand is 21 nucleotides in length. In another embodiment, the oligonucleotide of the application comprises at least one or at least two single stranded nucleotide overhang. In a particular embodiment, the oligonucleotide of the application is at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 97%, at least about 99% complementary or fully complementary (100% complementary) to an equal length portion of a target region selected from the group consisting of SEQ ID No. 2-4, 6-15, 17-18, 20-21, 23-25, 27-29, 31-41, 43- 45, 47-49, 51-57, 70-81, and 83-92.

In a most particular embodiment, the oligonucleotide of current application comprising a contiguous nucleotide sequence of at least 10 contiguous nucleotides in length that shows at least 90% sequence identity to any of SEQ. ID No. 172-249.

In one embodiment, the oligonucleotide of the application comprises one or more internucleoside linkage and/or one or more 2' sugar modified nucleosides, more particularly the internucleoside linkage is a phosphorothioate internucleoside linkage and/or the 2' sugar modified nucleoside is selected from the group consisting of 2'-O-methyl-, 2'-O-methoxyethyl-, 2'-O-alkyl-, 2' -alkoxy, 2' -amino-, 2'-fluoro- and LNA nucleosides. Even more particularly, all oligonucleosides are modified with a phosphorothioate internucleoside linkage or with a 2'-O-methyl group.

Also provided is an antisense oligonucleotide or RNAi molecule capable of reducing the level of synaptogyrin-3 mRNA, synaptogyrin-3 protein, synaptogyrin-3 activity, or a combination thereof in a cell by least 15% compared to a control situation in the absence of said antisense or RNAi molecule, wherein the antisense oligonucleotide or RNAi molecule nucleic acid sequence targets a subsequence of an mRNA encoding synaptogyrin-3 selected from the group consisting of SEQ ID NO. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 69, 82 and 93.

The oligonucleotides of the application, including the antisense oligonucleotides and RNAi molecules herein disclosed, are provided a therapeutic, more particularly to treat or reduce the symptoms of tauopathies. Therefore, pharmaceutical composition comprising the oligonucleotides of the application are provided as well as methods to treat tauopathies in a subject in need thereof, wherein the methods comprising the step of administering any of the antisense oligonucleotide and RNAi molecules herein provided. The oligonucleotide of the applications are also provided for use as a medicament, more particularly for use in treating or inhibiting progression of a tauopathic disorder or for use in treating or inhibiting a symptom of a tauopathic disorder.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 illustrates the structure of an exemplary siRNA molecule of the application. The molecule follows a 21/19 bp structure, where the sense strand is 19 nucleotides and the antisense strand is 21 nucleotides with 2 nucleotides overhanging on the 3' end. The strands are 2'0Me (green)/2"F (blue) modified. The red bar indicates a phosphorothioate modification, and "N" is complementary to the target mRNA. Figure 2 A-B show Modification scheme 1 and 2 respectively and illustrate alternative architectures of the oligonucleotides of the present disclosure.

Figure 3 shows the different target regions in the synaptogyrin-3 mRNA transcript that have been identified herein, their start (5') and end (3') position according to SEQ ID No. 1, their sequence, as well as the sequences of the sense and antisense strands of the siRNA molecules herein disclosed.

Figure 4 shows the extended target regions within the synaptogyrin-3 mRNA transcript.

DETAILED DESCRIPTION OF THE INVENTION

In one aspect, the current invention relates to oligonucleotides ("oligonucleotides of the present disclosure") that specifically bind to Synaptogyrin-3 RNA and reduce the expression of Synaptogyrin-3, e.g. through antisense or RNAi technology. In some aspect, the oligonucleotides of the present disclosure reduce Synaptogyrin-3 expression levels, Synaptopgyrin-3 activity (e.g., dopamine transporter activity), Synaptogyrin-3-mediated exocytosis, or a combination thereof. In some aspects, the oligonucleotide of the present disclosure is 10 to 50, 10 to 40, or of 10 to 30 nucleotides in length, and comprises a contiguous nucleotide sequence of at least 10 contiguous nucleotides in length which are at least 90% complementary to an equal portion of a target region within the Synaptogyrin-3 transcript as depicted in SEQ. ID No. 1. In some aspects, the target region within SEQ ID No. 1 is between nucleobase 205 and 265, between nucleobase 255 and 348, between nucleobase 338 and 387, between nucleobase 369 and 433, between nucleobase 422 and 531, between nucleobase 603 and 656, between nucleobase 641 and 714, between nucleobase 717 and 768, between nucleobase 1150 and 1600, between nucleobase 1743 and 1868, or between nucleobase 1865 and 2026 of SEQ ID No. 1, wherein the endpoints are included.

In some aspects, the oligonucleotide of the present disclosure is 10 to 50, 10 to 40, or of 10 to 30 nucleotides in length, and comprises a contiguous nucleotide sequence of at least 10 contiguous nucleotides in length which are at least 90% complementary to an equal length portion of a target region within human Synaptogyrin-3, wherein the target region is selected from the list consisting of SEQ ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 82 and 93.

In some aspects, the oligonucleotide of the present disclosure is complementary (full or partially complementary) to a target region of Synaptogyrin-3 selected from the group consisting of SEQ ID No. 2-4, SEQ ID No. 6-15, SEQ ID No. 17-18, SEQ ID No. 20-21, SEQ ID No. 23-25, SEQ ID No. 27-29, SEQ ID No. 31-41, SEQ ID No. 43-45, SEQ ID No. 47-49, SEQ ID No. 51-57, SEQ ID No. 70-81 and SEQ ID No. 83- 92.

In some aspects, the oligonucleotide of the application comprises or consists of 14, 15, 16, 17, 18, 19, 20, 21, 22 or 23 nucleotides in length. In other aspects, the oligonucleotide of the application comprises or consists of 19, 20, 21 or more nucleotides in length and comprises or consists of the sequence selected from the list consisting of SEQ. ID No. 94-249, more particularly of SEQ ID No. 172-249.

The present disclosure also provides methods of treatment comprising the administration of the oligonucleotides of the present disclosure, or a combination thereof, to a subject in need thereof. Also provides are pharmaceutical compositions, pharmaceutical formulations, and kits and articles of manufacture comprising the oligonucleotides of the present disclosure. Also provided are methods of manufacture of the oligonucleotides of the present disclosure.

Definitions

In order that the present description can be more readily understood, certain terms are first defined. Additional definitions are set forth throughout the detailed description. The present invention is described with respect to particular embodiments and with reference to certain drawings but the invention is not limited thereto but only by the claims. Any reference signs in the claims shall not be construed as limiting the scope. The drawings described are only schematic and are non-limiting. In the drawings, the size of some of the elements may be exaggerated and not drawn on scale for illustrative purposes.

It is to be noted that the term "a" or "an" entity refers to one or more of that entity; for example, "a nucleotide sequence", is understood to represent one or more nucleotide sequences. As such, the terms "a" (or "an"), "one or more" and "at least one" can be used interchangeably herein.

Furthermore, "and/or" where used herein is to be taken as specific disclosure of each of the two specified features or components with or without the other. Thus, the term "and/or" as used in a phrase such as "A and/or B" herein is intended to include "A and B", "A or B", "A" (alone), and "B" (alone). Likewise, the term "and/or" as used in a phrase such as "A, B, and/or C" is intended to encompass each of the following aspects: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

Where an indefinite or definite article is used when referring to a singular noun e.g. "a" or "an", "the", this includes a plural of that noun unless something else is specifically stated. Furthermore, the terms first, second, third and the like in the description and in the claims, are used for distinguishing between similar elements and not necessarily for describing a sequential or chronological order. It is to be understood that the terms so used are interchangeable under appropriate circumstances and that the embodiments of the invention described herein are capable of operation in other sequences than described or illustrated herein. It is understood that wherever aspects are described herein with the language "comprising", otherwise analogous aspects described in terms of "consisting of" and/or "consisting essentially of" are also provided. Where the term "comprising" is used in the present description and claims, it does not exclude other elements or steps. Unless specifically defined herein, all terms used herein have the same meaning as they would to one skilled in the art of the present invention.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure is related. For example, the Concise Dictionary of Biomedicine and Molecular Biology, Juo, Pei-Show, 2nd ed., 2002, CRC Press; The Dictionary of Cell and Molecular Biology, 3rd ed., 1999, Academic Press; and the Oxford Dictionary of Biochemistry and Molecular Biology, Revised, 2000, Oxford University Press, provide one of skill with a general dictionary of many of the terms used in this disclosure. Practitioners are particularly directed to Sambrook et al., Molecular Cloning: A Laboratory Manual, 4th ed., Cold Spring Harbor Press, Plainsview, New York (2012); and Ausubel et al., current Protocols in Molecular Biology (Supplement 100), John Wiley & Sons, New York (2012), for definitions and terms of the art. The definitions provided herein should not be construed to have a scope less than understood by a person of ordinary skill in the art.

Units, prefixes, and symbols are denoted in their Systeme International de Unites (SI) accepted form. Numeric ranges are inclusive of the numbers defining the range. Unless otherwise indicated, nucleotide sequences are written left to right in 5' to 3' orientation. Amino acid sequences are written left to right in amino to carboxy orientation. The headings provided herein are not limitations of the various aspects of the disclosure, which can be had by reference to the specification as a whole. Accordingly, the terms defined immediately below are more fully defined by reference to the specification in its entirety.

The term "about" is used herein to mean approximately, roughly, around, or in the regions of. When the term "about" is used in conjunction with a numerical range, it modifies that range by extending the boundaries above and below the numerical values set forth. In general, the term "about" can modify a numerical value above and below the stated value by a variance of, e.g., 10 percent, up or down (higher or lower). For example, if it is stated that an oligonucleotide of the present disclosure reduces expression the Syngr-3 transcript in a cell following administration of an oligonucleotide of the present disclosure by at least about 60%, it is implied that the Syngr-3 expression levels are reduced by a range of 50% to 70%.

The terms "reverse complement", "reverse complementary" and "reverse complementarity" as used herein are interchangeable with the terms "complement", "complementary" and "complementarity". The terms "identical" or percent "identity" in the context of two or more nucleic acids refer to two or more sequences that are the same or have a specified percentage of nucleotides or amino acid residues that are the same, when compared and aligned (introducing gaps, if necessary) for maximum correspondence, not considering any conservative amino acid substitutions as part of the sequence identity. The percent identity can be measured using sequence comparison software or algorithms or by visual inspection. Various algorithms and software are known in the art that can be used to obtain alignments of amino acid or nucleotide sequences.

The term "percent sequence identity" or "percent identity" between two polynucleotide or polypeptide sequences refers to the number of identical matched positions shared by the sequences over a comparison window, taking into account additions or deletions (i.e. gaps) that must be introduced for optimal alignment of the two sequences. A matched position is any position where an identical nucleotide or amino acid is presented in both the target and reference sequence. Gaps presented in the target sequence are not counted since gaps are not nucleotides or amino acids. Likewise, gaps presented in the reference sequence are not counted since target sequence nucleotides or amino acids are counted, not nucleotides or amino acids from the reference sequence.

One such non-limiting example of a sequence alignment algorithm is the algorithm described in Karlin et al., 1990, Proc. Natl. Acad. Sci., 87:2264-2268, as modified in Karlin et al., 1993, Proc. Natl. Acad. Sci., 90:5873-5877, and incorporated into the NBLAST and XBLAST programs (Altschul et al., 1991, Nucleic Acids Res., 25:3389-3402). In certain aspects, Gapped BLAST can be used as described in Altschul et al., 1997, Nucleic Acids Res. 25:3389-3402. BLAST-2, WU-BLAST-2 (Altschul et al., 1996, Methods in Enzymology, 266:460-480), ALIGN, ALIGN-2 (Genentech, South San Francisco, California) or Megalign (DNASTAR) are additional publicly available software programs that can be used to align sequences. In certain aspects, the percent identity between two nucleotide sequences is determined using the GAP program in the GCG software package (e.g., using a NWSgapdna.CMP matrix and a gap weight of 40, 50, 60, 70, or 90 and a length weight of 1, 2, 3, 4, 5, or 6). In certain alternative aspects, the GAP program in the GCG software package, which incorporates the algorithm of Needleman and Wunsch (J. Mol. Biol. (48):444-453 (1970)) can be used to determine the percent identity between two amino acid sequences (e.g., using either a BLOSUM 62 matrix or a PAM 250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5). Alternatively, in certain aspects, the percent identity between nucleotide or amino acid sequences is determined using the algorithm of Myers and Miller (CABIOS, 4:11-17 (1989)). For example, the percent identity can be determined using the ALIGN program (version 2.0) and using a PAM 120 with residue table, a gap length penalty of 12 and a gap penalty of 4. One skilled in the art can determine appropriate parameters for maximal alignment by particular alignment software. In certain aspects, the default parameters of the alignment software are used. One skilled in the art will appreciate that the generation of a sequence alignment for the calculation of a percent sequence identity is not limited to binary sequence-sequence comparisons exclusively driven by primary sequence data. Sequence alignments can be derived from multiple sequence alignments. One suitable program to generate multiple sequence alignments is ClustalW2, available from www.clustal.org. Another suitable program is MUSCLE, available from www.drive5.com/muscle/. ClustalW2 and MUSCLE are alternatively available, e.g., from the EBI (European Bioinformatics Institute). In certain aspects, the percentage identity "X" of a first nucleotide sequence to a second nucleotide sequence is calculated as 100 x (Y/Z), where Y is the number of amino acid residues scored as identical matches in the alignment of the first and second sequences (as aligned by visual inspection or a particular sequence alignment program) and Z is the total number of residues in the second sequence. If the length of a first sequence is longer than the second sequence, the percent identity of the first sequence to the second sequence will be higher than the percent identity of the second sequence to the first sequence. Different regions within a single polynucleotide target sequence that align with a polynucleotide reference sequence can each have their own percent sequence identity. It is noted that the percent sequence identity value is rounded to the nearest tenth. For example, 80.11, 80.12, 80.13, and 80.14 are rounded down to 80.1, while 80.15, 80.16, 80.17, 80.18, and 80.19 are rounded up to 80.2. It also is noted that the length value will always be an integer.

As used in the present disclosure, the terms "nucleic acid molecule of the invention" and "oligonucleotide of the present disclosure" and grammatical variants thereof are used interchangeably. The term "defined by SEQ ID No. X" as used herein refers to a biological sequence consisting of the sequence of nucleotides given in the SEQ. ID No. X. SEQ ID No. X is interchangeable with SEQ ID NO: X. When the present application refers to "a group consisting of SEQ ID No. 2-4", this is identical to a group consisting of SEQ ID No. 2, SEQ ID No. 3 and SEQ ID No. 4.

The target nucleic acid of the invention is a nucleic acid, e.g., an mRNA, encoding Synaptogyrin-3, more particularly human Synaptogyrin-3. "Synaptogyrin3", "Synaptogyrin3", "synaptogyrin-3", "synaptogyrin- 3", "Syngr3", "Syngr-3", "SYNGR3" or "SYNGR-3" are interchangeably used and refer herein to Synaptogyrin-3 transcript if not otherwise specified. The human nucleic acid sequence of Synaptogyrin-3 (hSyngr-3) is set forth in SEQ ID NO: 1; however also within the scope of the invention are nucleic acid sequence variants of Synaptogyin-3 as may exist due to allelic variation, e.g., a mRNA encoding a Synaptogyrin-3 allelic variant. Such variations are defined herein as "allelic variants of SEQ ID NO: 1". The term "allelic variants" refer to one of several alternate forms of a gene occupying a given locus on a chromosome of an organism (Genes II, Lewin, B., ed., John Wiley & Sons, New York (1985)). These allelic variants can vary at either the polynucleotide and/or polypeptide level and are included in the present disclosure. Alternatively, non-naturally occurring variants can be produced by mutagenesis techniques or by direct synthesis. In some aspects, the synaptogyrin-3 variant is a splice variant. In some aspects, the synaptogyrin-3 variant is a post-translationally modified variant. In some aspects, the synaptogyrin- 3 variant is a mutant synaptogyrin-3, e.g., a mutant comprising at least one nucleotide point mutation, deletion, or insertion. In some aspects, the mutation is a silent mutation. In some aspects, the synaptogyrin-3 variant is a mutant protein comprising at least one amino acid substitution, deletion, or insertion. In some aspects, the synaptogyrin-3 variant is a loss of function variant. In some aspects, the synaptogyrin-3 variant is a gain of function variant.

"Specific to synaptogyrin-3" as used herein is referring to the fact that the nucleic acid molecule or oligonucleotide of the invention is acting at the level of synaptogyrin-3 and not at the level of another transcript. Specificity can be ascertained by e.g. determining the expression level of closely related RNA sequences.

The term "statistically significantly" different is well known by the person skilled in the art. Statistical significance plays a pivotal role in statistical hypothesis testing. It is used to determine whether the null hypothesis should be rejected or retained. The null hypothesis is the default assumption that nothing happened or changed, hence that there is no difference for example in the synaptogyrin-3 transcript level in the presence of an antisense or RNAi molecule compared to the synaptogyrin-3 transcript level in the absence of said antisense or RNAi molecule. For the null hypothesis to be rejected, an observed result has to be statistically significant, i.e. the observed p-value is less than the pre-specified significance level a. The p-value of a result, p, is the probability of obtaining a result at least as extreme, given that the null hypothesis were true. In one embodiment, a is 0.05. In a more particular embodiment, a is 0.01. In an even more particular embodiment, a is 0.001.

Nucleic acid molecules inhibiting the expression of Synaptogyrin-3

In a first aspect, the application discloses nucleic acid molecules, more particularly oligonucleotides, that comprise a sequence complementary (fully or partially) to a region of an mRNA encoding human Synaptogyrin-3, e.g. a mature mRNA as depicted in SEQ ID No. 1, or an mRNA encoding an allelic variant or isoforms thereof (e.g., any of the variants and isoform disclosed in the UniProtKB/Swiss-Prot 043761 entry).

SEQ. ID No. 1 represent the mature mRNA sequence of the human Synaptogyrin-3 gene (Gene ID: 9143; Sequence ID: NM_004209.6). The mRNA is 2026 bp long, comprises 4 exons and encodes the human SYNAPTOGYRIN-3 protein (UniProtKB/Swiss-Prot: 043761). In some aspects the target mRNA is a pre- mRNA or a splice variant of a pre-mRNA. In some aspects, the target sequence comprises an exon, an intron, or a combination thereof.

The Synaptogyrin-3 coding sequence is underlined in the sequence below. " |" indicates the location of an exon junction.

SEQ ID No. 1 (Synaptogyrin-3 mRNA)

GAGGCGGCAGCGGCTGCAGCGTTGGTAGCATCAGCATCAGCATCAGCGGCAGCGGCAGCGGCCTCGGGCGGGGCCGGCC GGACGGACAGGCGGACAGAAGGCGCCAGGGGCGCGCGTCCCGCCCGGGCCGGCCATGGAGGGCGCCTCCTTCGGCGCGG GCCGCGCAGGGGCCGCCCTGGACCCCGTGAGCTTTGCGCGGCGGCCCCAGACCCTGCTCCGGGTCGCGTCCTGG I GTGTTCT CCATCGCCGTCTTCGGGCCCATCGTCAACGAGGGCTACGTGAACACCGACAGCGGCCCCGAGCTGCGCTGCGTGTTCAACGG GAACGCGGGCGCCTGCCGCTTCGGCGTCGCGCTGGGCCTCGGAGCCTTCCTCGCCTGCGCCGCCTTCCTGCTGCTCGATGTG CGCTTCCAGCAAATCAGCAGCGTCCGCGACCGCCGGCGCGCGGTGTTGCTGGACCTGGGCTTCTCAG | GACTCTGGTCCTTC CTGTGGTTCGTGGGCTTCTGCTTCCTCACCAATCAGTGGCAGCGCACGGCGCCAGGGCCGGCCACGACGCAGGCGGGGGAC GCGGCGCGGGCCGCCATCGCCTTCAGCTTCTTCTCCATCCTCAGCTGG | GTGGCGCTCACCGTGAAGGCCCTGCAGCGGTTCC GCCTGGGCACCGACATGTCACTCTTCGCCACCGAACAGCTGAGCACCGGGGCGAGCCAGGCCTACCCCGGCTATCCGGTGG GCAGCGGCGTGGAGGGCACCGAGACCTACCAGAGCCCGCCCTTCACCGAGACCCTGGACACCAGCCCCAAAGGGTACCAG GTGCCCGCCTACTAGCGGCTGGCAGGCACAGACCAGGGCTCCAAGGCCACCCCACCAACGCAGGCCCCAGGGTCTCCGGGA CCTCCCTTGGGTCCTTCCAGCTCAGTGCCGCGGACAGAGTAGGTGGCCGCTTTGCGCCATCCGGGGCCAAGAGGGGGTGGA CCCGCGTGTCTGGGCTGCCCCTGCCAAGTTCCCCCAGTCCCTCAGCACCTGGCCCCAGGACTGAGGTCCTGAGAAGGGGATA GCACTGCCCAGGACGTGTGTCCCTAGCCTGGAATGGACTGGCCTGGGGAAGGCTTTCCCCTCTTGGGCCACACCTGCTCACT CTGGGGTTGGGGGTCCAGCTGCCCTCTACGATCAGGTGCAGGGGCTGCCCAGGACAAAGCGGGGGCAGGGGAAAGACACC ACCCTCGCCCCAAGACTGGGGATCCTGGCCACTGTTCCCATCCCATGTCCCTGTGGGTAGTGACTGTCTCGTTTCTGTCATGG TGGTGCGTCCCGTCCGGAGCCACTCTCCACTTTCTCTCACAGGCTGCTAGAACAGCCCAGCCCTGTCAGTGTTGTGATCATGG TCCAGTCTTCGGGTTTCACCTCCTAGTACTCCACAAGCTGCTCCTCTCTCTGTGGCCCCGGCCCCTGCCCAGGTGTGGGTGGTT CTGGCCAGGAAGGCACAAGGTAGCTGTGGGCCAAGACACCAGCCCTGTCCTAGCCCTTCAGTAAGACCTTGCCAGGAGAGG AGAAGGATGCCTGGGTGCCAGGCAAGACAAGCCCCTCAGCAGGAGAGAGGCCCAGAGGCTCCAGCTGGCCACCGTGCCCC ACAAGATGGCCCCTGTGTGGTTCCCTTTACCTTGGCTTCCTGGCCCAGTCCCTGCCTCTCCACCTGCACCCTGCTTCCTGGCCC AGTCCCAGGTTGGAGTCCCTCTGCATAGCTGACTACTCATGCATTGCTCAAAGCTGGCTTTTCACATTAAGTCAACACCAAAC GTGGTTGCCACATTTCATCAGACAGACACCTCCCTCTGGAGATGCAGTTGAGTGACAACCTTGTTACATTGTAGCCTAGACCA

ATTCTGTGTGGATATTTAAGTGAACATGTTTACAATTTTTGTATATATCACTCTCTCCCTCTCCTGAAAGACCAGAGATTGTGT ATTTTCAGTGTCCCATGTTCCGACTGCACCTTCTTTACAATAAAGACTGTAACTGAGCTGACTGTGA

In some aspects, the nucleic acid molecule or the oligonucleotide of the present disclosure (e.g. ASO, siRNA, shRNA) comprises an oligonucleotide of between 10 and 50 nucleotides in length.

In some aspects, the nucleic acid molecule or the oligonucleotide of the present disclosure comprises of or consists of 1 oligonucleotide (e.g., an ASO oligomer or a shRNA). In some aspects, the nucleic acid molecule or the oligonucleotide of the present disclosure comprises or consists of 2 oligonucleotides (e.g., a siRNA). In some aspects, the 2 oligonucleotides are a sense oligonucleotide and an antisense oligonucleotide. In some aspects, the sense oligonucleotide and the antisense oligonucleotide are connected by a loop. In some aspects, a nucleic acid molecule of the present disclosure comprises or consists of a sequence from about 8 to about 70 contiguous nucleotides in length disclosed herein, a sequence from about 10 to about 60 contiguous nucleotides in length disclosed herein, a sequence from about 12 to about 50 contiguous nucleotides in length disclosed herein, a sequence from about 8 to about 40 nucleotides in length, a sequence from about 10 to about 35 contiguous nucleotides in length disclosed herein, a sequence from about 12 to about 30 contiguous nucleotides in length disclosed herein, a sequence from about 14 to about 28 contiguous nucleotides in length disclosed herein, a sequence from about 16 to about 25 nucleotides in length disclosed herein, a sequence from about 17 to about 24 nucleotides in length disclosed herein, or a sequence from about 18 to about 23 contiguous nucleotides in length disclosed herein. In some aspects, the nucleic acid molecule of the invention is 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, or 60 nucleotides in length. In one embodiment, "nucleotides in length" refers to "contiguous nucleotides in length".

"Nucleotides" as used herein refer to the building blocks of oligonucleotides and polynucleotides, and for the purposes of the present invention include both naturally occurring and non-naturally occurring nucleotides. In nature, nucleotides, such as DNA and RNA nucleotides comprise a ribose sugar moiety, a nucleobase moiety and one or more phosphate groups (which are absent in nucleosides). A nucleotide without a phosphate group is called a "nucleoside" and is thus a compound comprising a nucleobase moiety and a sugar moiety. As used herein, "nucleobase" means a group of atoms that can be linked to a sugar moiety to create a nucleoside that is capable of incorporation into an oligonucleotide, and wherein the group of atoms is capable of bonding with a complementary naturally occurring nucleobase of another oligonucleotide or nucleic acid. Naturally occurring nucleobases of RNA or DNA comprise the purine bases adenine (A) and guanine (G), and the pyrimidine bases thymine (T), cytosine (C) and uracil (U).

The term "contiguous nucleotides" or "contiguous nucleotide sequence" as used herein refers to the uninterrupted region of the oligonucleotide which is complementary to the target nucleic acid. "Contiguous" as used herein means next or together in sequence, hence the contiguous nucleotides are linked nucleotides (i.e. no additional nucleosides are present between those that are linked). The target nucleic acid of the invention is Synaptogyrin-3, more particularly human Synaptogyrin-3.

The terms "oligomer" or "oligonucleotide" in the context of the present disclosure are used interchangeably and refer to a molecule formed by covalent linkage of two or more nucleotides. Herein, a single nucleotide (unit) can also be referred to as a monomer or unit. In some aspects, the present disclosure provides a derivative of an oligonucleotide of the present disclosure which is a conjugate, e.g., a GalNAc conjugate. The term "derivative" as used herein refers to a chemical compound related structurally to a compound disclosed herein (e.g., an oligonucleotide of the present disclosure), e.g., having the same carbon skeleton, but chemically modified to introduce, e.g., a side chain or group, in one or more positions, and wherein the derivative possesses a biological activity (e.g., the capacity to reduce Syngr3 expression) that is substantially similar to a biological activity of the entity or molecule it is a derivative.

The term, "complementary" means that two sequences are complementary when the sequence of one can bind to the sequence of the other in an anti-parallel sense wherein the 3'-end of each sequence binds to the 5'-end of the other sequence and each A, T(U), G, and C of one sequence is then aligned with a T(U), A, C, and G, respectively, of the other sequence. Normally, the complementary sequence of the oligonucleotide has at least 90%, preferably 95%, most preferably 100% complementarity to a defined sequence.

In determining the degree of "complementarity" between oligonucleotides of the disclosure (or regions thereof) and the target region, such as those disclosed herein, the degree of "complementarity" (also, "homology" or "identity") is expressed as the percentage identity (or percentage homology) between the sequence of the oligonucleotide (or region thereof) and the sequence of the target region (or the reverse complement of the target region) that best aligns therewith. The percentage is calculated by counting the number of aligned bases that are identical between the two sequences, dividing by the total number of contiguous monomers (e.g. nucleotides) in the oligomer (e.g. oligonucleotide), and multiplying by 100. In such a comparison, if gaps exist, it is preferable that such gaps are merely mismatches rather than areas where the number of monomers within the gap differs between the oligomer of the disclosure and the target region.

In some embodiments, the nucleic acid molecule described above or the contiguous nucleotide sequence thereof comprises or consists of less than 60 nucleotides, less than 59 nucleotides, less than 58 nucleotides, less than 57 nucleotides, less than 56 nucleotides, less than 55 nucleotides, less than 54 nucleotides, less than 53 nucleotides, less than 52 nucleotides, less than 51 nucleotides, less than 50 nucleotides, less than 49 nucleotides, less than 48 nucleotides, less than 47 nucleotides, less than 46 nucleotides, less than 45 nucleotides, less than 44 nucleotides, less than 43 nucleotides, less than 42 nucleotides, less than 41 nucleotides, less than 40 nucleotides, less than 39 nucleotides, less than 38 nucleotides, less than 37 nucleotides, less than 36 nucleotides, less than 35 nucleotides, less than 34 nucleotides, less than 33 nucleotides, less than 32 nucleotides, less than 31 nucleotides, less than 30 nucleotides, less than 29 nucleotides, less than 28 nucleotides, less than 27 nucleotides, less than 26 nucleotides, less than 25 nucleotides, less than 24 nucleotides, less than 23 nucleotides, less than 22 nucleotides, less than 21 nucleotides, less than 20 nucleotides, less than 19 nucleotides, less than 18 nucleotides, less than 17 nucleotides, less than 16 nucleotides, less than 15 nucleotides, less than 14 nucleotides, less than 13 nucleotides, or less than 12 nucleotides.

It is to be understood that any range given in current application includes the range endpoints. Accordingly, if a nucleic acid molecule is said to include from 10 to 30 nucleotides, both 10 and 30 nucleotides are included.

In some embodiments, the contiguous nucleotide sequence comprises or consists of 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 1 , 28, 29 or 30 contiguous nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 14 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 15 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 16 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 17 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 18 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 19 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 20 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 21 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 22 nucleotides in length. In some aspects, the nucleic acid molecule of the invention is 24 nucleotides in length.

The nucleic acid molecule(s) is typically for modulating the expression of Synaptogyrin-3 as target nucleic acid in a mammal. In some embodiments the nucleic acid molecule(s), such as siRNAs, shRNAs or antisense oligonucleotides, is typically for inhibiting the expression of a target nucleic acid. More particularly, the oligonucleotides of the application are provided as being capable of reducing the level of Synaptogyrin-3 mRNA transcript (and thus indirectly SYNGR3 protein) in a cell, wherein the reduction is determined by comparison to a control situation, i.e. the level of Synaptogyrin-3 mRNA transcript in the same cell or same cell type grown in the same conditions but in the absence of the oligonucleotide of the application.

The oligonucleotides described in the application are partially or fully complementary to an equal length portion of a target region within the Synaptogyrin-3 as depicted in SEQ. ID No. 1 or allelic variants thereof. The specific target regions will be described in detail below. In another embodiment, the oligonucleotide of the invention comprises a contiguous nucleotide sequence which is complementary to one of said specific target regions, and may, in some embodiments further comprise one or more additional nucleotides, such as 1-30, such as 1-20, such as 1-10, such as 1, 2, 3, 4 or 5 further nucleotides in addition to the contiguous nucleotide sequence. In some embodiments the additional nucleotides are complementary to the contiguous nucleotide sequence and are capable of forming a stem loop (hairpin) structure by hybridizing to the contiguous nucleotide sequence. In some embodiments the additional nucleotides are 1 to 5 phosphodiester linked nucleotides. In some embodiments, all the nucleotides of the oligonucleotide form the contiguous nucleotide sequence.

In yet another embodiment, the oligonucleotide of the invention may be or comprise an antisense oligonucleotide (ASO) or may be another oligomeric nucleic acid molecule such as a CRISPR RNA, a siRNA, shRNA, an aptamer or a ribozyme. In a particular embodiment the oligonucleotide of the invention is a RNAi molecule or RNAi agent, more particularly a siRNA, di-siRNA, a shRNA or a miRNA.

The term "RNAi agent" or "RNA interference (RNAi) molecule" refers to any molecule inhibiting RNA expression or translation via the RNA reducing silencing complex (RISC) in a cell's cytoplasm, where the RNAi molecule interacts with the catalytic RISC component argonaut. A small interfering RNA (siRNA) is typically a double-stranded RNA complex comprising a passenger (sense) and a guide (antisense) oligonucleotide (strand), which when administered to a cell, results in the incorporation of the guide (antisense) strand into the RISC complex (si R ISC) resulting in the RISC associated inhibition of translation or degradation of complementary RNA target nucleic acids in the cell. The sense strand is also referred to as the passenger strand, and the antisense strand as the guide strand. A small hairpin RNA (shRNA) is a single nucleic acid molecule which forms a stem loop (hairpin) structure that is able to degrade mRNA via RISC. RNAi nucleic acid molecules may be synthesized chemically (typical for siRNA complexes) or by in vitro transcription, or expressed from a vector. shRNA molecules are generally between 40 and 70 nucleotides in length, such as between 45 and 65 nucleotides in length, such as 50 and 60 nucleotides in length, and interacts with the endonuclease known as Dicer which is believed to processes dsRNA into 19-23 base pair short interfering RNAs with characteristic two base 3' overhangs which are then incorporated into an RNA-induced silencing complex (RISC). Typically, the guide (antisense) strand of an siRNA (or antisense region of a shRNA) is 17-25 nucleotide in length, such as 19-23 nucleotides in length and complementary to the target nucleic acid or target sequence. In an siRNA complex, the guide (antisense) strand and passenger (sense) strand form a double stranded duplex, which may comprise 3' terminal overhangs of e.g. 1- 3 nucleotides (resembles the product produced by Dicer), or may be blunt ended (no overhang at one or both ends of the duplex). It will be recognized that RNAi may be mediated by longer dsRNA substrates which are processed into siRNAs within the cell (a process which is thought to involve the dsRNA endonuclease DICER), such as miRNAs. In another particular embodiment, the nucleic acid molecule of the invention or the oligonucleotide of the disclosure is an antisense oligonucleotide (ASO), such as single stranded antisense oligonucleotide, such as a high affinity modified antisense oligonucleotide interacting with RNase H.

The term "antisense oligonucleotide" or "ASO" as used herein is defined as an oligonucleotide capable of modulating expression of a target gene by hybridizing to a target nucleic acid, in particular to a contiguous sequence on a target nucleic acid. The antisense oligonucleotides are not essentially double stranded and are therefore not siRNAs or shRNAs. Preferably, the antisense oligonucleotides of the present invention are single stranded. It is understood that single stranded oligonucleotides of the present invention can form hairpins or intermolecular duplex structures (duplex between two molecules of the same oligonucleotide), as long as the degree of intra or inter self-complementarity is less than 50% across of the full length of the oligonucleotide.

In a further particular embodiment, the single stranded antisense oligonucleotide of the invention does not contain RNA nucleosides, since this will decrease nuclease resistance. More particularly, the antisense oligonucleotide of the invention comprises one or more modified nucleosides or nucleotides, such as 2' sugar modified nucleosides. Furthermore, it is advantageous that the nucleosides which are not modified are DNA nucleosides.

In one embodiment, the oligonucleotide, e.g. the therapeutic antisense oligonucleotide, shRNA or siRNA, of the invention comprises one or more internucleoside linkages modified from the natural phosphodiester, such one or more modified internucleoside linkages that is for example more resistant to nuclease attack. The term "modified internucleoside linkage" is defined as generally understood by the skilled person as linkages other than phosphodiester (PO) linkages, that covalently couples two nucleosides together. Increased resistance of the oligonucleotide towards nucleases compared to a phosphodiester linkage is particular advantage for therapeutic oligonucleotides. Nuclease resistance may be determined by incubating the oligonucleotide in blood serum or by using a nuclease resistance assay (e.g. snake venom phosphodiesterase (SVPD)), both are well known in the art. Internucleoside linkages which are capable of enhancing the nuclease resistance of an oligonucleotide are referred to as nuclease resistant internucleoside linkages. In some embodiments at least 50% of the internucleoside linkages in the oligonucleotide, or contiguous nucleotide sequence thereof, are modified, such as at least 60%, such as at least 70%, such as at least 80 or such as at least 90% of the internucleoside linkages in the oligonucleotide, or contiguous nucleotide sequence thereof, are nuclease resistant internucleoside linkages. In some embodiments all of the internucleoside linkages of the oligonucleotide, or contiguous nucleotide sequence thereof, are nuclease resistant internucleoside linkages. It will be recognized that, in some embodiments the nucleosides which link the oligonucleotide of the invention to a non- nucleotide functional group, such as a conjugate, may be phosphodiester. In a particular embodiment, the modified internucleoside linkage is phosphorothioate.

Phosphorothioate internucleoside linkages are particularly useful due to nuclease resistance, beneficial pharmacokinetics and ease of manufacture. In some embodiments at least 50% of the internucleoside linkages in the oligonucleotide, or contiguous nucleotide sequence thereof, are phosphorothioate, such as at least 60%, such as at least 70%, such as at least 80% or such as at least 90% of the internucleoside linkages in the oligonucleotide, or contiguous nucleotide sequence thereof, are phosphorothioate. In some embodiments all of the internucleoside linkages of the oligonucleotide, or contiguous nucleotide sequence thereof, are phosphorothioate. The use of fully phosphorothioate modified oligonucleotides or contiguous nucleotide sequences is often used in antisense oligonucleotides, although in siRNAs partial phosphorothioate modifications may be preferred as fully phosphorothioate modifications have been reported to limit RNAi activity, particularly when used in the guide (antisense) strand. Phosphorothioate modifications may be incorporated into the 5' and 3' ends of an antisense strand of a siRNA without unduly limiting RNAi activity.

Nuclease resistant linkages, such as phosphorothioate linkages, are particularly useful in oligonucleotide regions capable of recruiting nuclease when forming a duplex with the target nucleic acid, such as region G for gapmers. Phosphorothioate linkages may, however, also be useful in non-nuclease recruiting regions and/or affinity enhancing regions such as regions F and F' for gapmers. Gapmer oligonucleotides may, in some embodiments comprise one or more phosphodiester linkages in region F or F', or both region F and F', which the internucleoside linkage in region G may be fully phosphorothioate. In particular embodiments, all the internucleoside linkages in the contiguous nucleotide sequence of the antisense oligonucleotide are phosphorothioate linkages.

In other embodiments, antisense oligonucleotide may comprise other internucleoside linkages (other than phosphodiester and phosphorothioate), for example alkyl phosphonate / methyl phosphonate internucleosides.

In some embodiments, the RNAi molecules of the invention comprise one or more phosphorothioate internucleoside linkages. In RNAi molecules phosphorothioate internucleoside linkages may reduce the nuclease cleavage in RICS and it is therefore advantageous that not all internucleoside linkages are modified. Phosphorothioate internucleoside linkages can advantageously be place in the 3' and/or 5' end of the RNAi molecule, in particular in part of the molecule that is not complementary to the target nucleic acid (e.g. the sense strand or passenger strand in an siRNA molecule). The region of the RNAi molecule that is complementary to the target nucleic acid (e.g. the antisense or guide strand in a siRNA molecule) may however also be modified in the first 2 to 3 internucleoside linkages in the 3' and/or 5' terminal. In other embodiments, the oligonucleotides of the invention may be chemically modified by incorporating high affinity nucleosides such as 2' sugar modified nucleosides, such as 2' -4' bicyclic ribose modified nucleosides, including LNA and cET or 2' substituted modifications like of 2'-O-alkyl-RNA, 2'-O- methyl-RNA, 2'-alkoxy-RNA, 2'-O-methoxyethyl-RNA (MOE), 2'-amino-DNA, 2'-fluoro-DNA, arabino nucleic acid (ANA), 2'-fluoro-ANA. See for example WO 2002/044321 which discloses 2'-O-Methyl modified siRNAs, W02004083430 which discloses the use of LNA nucleosides in siRNA complexes, known as siLNAs, and W02007107162 which discloses the use of discontinuous passenger strands in siRNA such as siLNA complexes.

In a particular embodiment, the oligonucleotide of the invention comprises a 2' sugar modified nucleoside selected from the list consisting of 2'-O-methyl (2'-0Me), 2'-O-methoxyethyl (2'-MOE) and 2'- Fluoro (2'-F).

In other embodiments, the oligonucleotides of the invention may comprise one or more of the above described chemically modified sugar nucleosides and may comprise one or more of the above described phosphorothioate internucleoside linkages.

The skilled person is aware of how to design the oligonucleotides disclosed herein. siRNA and shRNA design programs are publicly available. Non-limiting examples are siDESIGN from ThermoScientific, siDirect (Naito et al), BLOCK-IT RNAi Designer from Invitrogen, siRNA Wizard from InvivoGen, shRNA design tool from Gene Link and shRNA design tool from transomic. Manufacturers of RNAi products also provide guidelines for designing siRNA/shRNA. siRNA sequences between 19-29 nucleotides (nt) are generally the most effective. Sequences longer than 30 nt can result in nonspecific silencing. Ideal sites to target include AA dinucleotides and the 19 nt 3' of them in the target mRNA sequence. Typically, siRNAs with 3' dUdU or dTdT dinucleotide overhangs are more effective. Other dinucleotide overhangs could maintain activity but GG overhangs should be avoided. Also to be avoided are siRNA designs with a 4-6 poly(T) tract (acting as a termination signal for RNA pol III), and the G/C content is advised to be between 35-55%. shRNAs should comprise sense and antisense sequences (advised to each be 19-21 nt in length) separated by loop structure, and a 3' AAAA overhang. Effective loop structures are suggested to be 3-9 nt in length. It is suggested to follow the sense-loop-antisense order in designing the shRNA cassette and to avoid 5' overhangs in the shRNA construct. Finally, several companies commercially offer premade siRNAs and shRNAs.

In some aspects, any of the oligonucleotides of present disclosure are provided, wherein the oligonucleotide is an RNAi molecule such as a siRNA, shRNA or di-siRNA comprising at least one nucleotide variant (e.g., an LNA unit). In some aspects, the oligonucleotide of the present disclosure further comprises at least one non-nucleotide or non-polynucleotide moiety covalently (e.g., a GalNac moiety) attached to said oligonucleotide directly or via a linker positioned between the contiguous nucleotide sequence and the non-nucleotide or non-polynucleotide moiety.

In some aspects, the present disclosure provides oligonucleotides of the present disclosure comprising 16 to 22 contiguous oligonucleotides in length comprising a contiguous sequence of 16 nucleotides in length which is 100% complementary to a human synaptogyrin-3 target sequence selected from the group consisting of SEQ ID No. 2-93, wherein oligonucleotide is an RNAi molecule such as a siRNA, shRNA or di-siRNA comprising at least one nucleotide variant (e.g., an LNA unit), and wherein the RNAi molecule targets the synaptogyrin-3 transcript as set forth in SEQ ID No. 1.

In some aspects, the oligonucleotide of the present disclosure comprises, consists, or consists essentially of an RNAi molecule binding to the human synaptogyrin-3 transcript as set forth in SEQ. ID No. 1, the RNAi molecule comprising one or more sequences selected from the group consisting of SEQ ID No. 94- 249, more particularly of SEQ ID No. 172-249.

In some aspects, the oligonucleotide of the present disclosure comprises an RNAi molecule comprising one or more sequences selected from the group consisting of SEQ ID No. 94-249, except for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 nucleobase substitutions. In some aspects, the oligonucleotide of the present disclosure comprises an RNAi molecule comprising a sequence which is at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% identical to a sequence selected from the group consisting of SEQ ID No. 94-249. In some aspects, the oligonucleotide of the present disclosure comprises an RNAi molecule comprising a sequence which is about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% identical to a sequence selected from the group consisting of SEQ ID No. 172-249. In some aspects, the oligonucleotide of the present disclosure comprises a sequence that overlaps with 9, 10, 11, 12, 13, 14, 15, or 16 nucleobase subsequence from a sequence selected from the group consisting of SEQ ID No. 172-249.

In some aspects, the oligonucleotide of the present disclosure comprises at least one non-cleavable internucleoside linkage, e.g., a phosphorothioate linkage. In some, aspects all the internucleoside linkages in an oligonucleotide of the present disclosure are non-cleavable, e.g., phosphorothioates linkages. In some aspects, the non-cleavable internucleoside linkages, e.g., a phosphorothioate linkages, are present only in the wing portions of a gapmer, e.g., the last 1, 2 or 3 linkages at the 5' end or the oligonucleotide, and the last 1, 2 or 3 linkages at the 5' end or the oligonucleotide. In some aspects, the oligonucleotide of the present disclosure comprises nucleotide analogues. In some aspects, the oligonucleotide of the present disclosure comprises affinity enhancing nucleotide analogues. In some aspects, the nucleotide analogues are sugar modified nucleotides, such as sugar modified nucleotides independently or dependently selected from the group consisting of 2'-O-alkyl-RNA units, 2'-0Me-RNA units, 2'-amino-DNA units, and 2'-fluoro-DNA units.

In some aspects, the oligonucleotide of the present disclosure is an siRNA, di-siRNA, shRNA, RNA duplex or the antisense strand from an RNA duplex. In some aspects, the oligonucleotide of the present disclosure comprises one or more locked nucleic acids (LNA). In some aspects, the LNA oligonucleotide comprises a wing on each side (5' and 3') of 2 to 4 nucleotide analogues, preferably LNA analogues. In some aspects, the oligonucleotide of the present disclosure can optionally comprise a further 1 to 6 nucleotides (e.g., one, two, three, four, five or six nucleotides), which can form or comprise a biocleavable nucleotide region, such as a phosphate nucleotide linker. In some aspects, the biocleavable nucleotide region is formed of a short stretch of nucleotides (e.g. 1, 2, 3, 4, 5 or 6 nucleotides) which are physiologically labile. This can be achieved by using phosphodiester linkages with DNA/RNA nucleosides, or if physiological liability can be maintained, other nucleoside can be used.

In some aspects, the LNA is oxy-LNA, thio-LNA, amino-5 LNA, 5'-methyl-LNA, ENA, cET, cMOE or a combination thereof. In some aspects, the LNA is an stereoisomer in the beta-D configuration or the alpha-L configuration. In some aspects, the oligonucleotide of the application comprises at least one cET unit. In some aspects, the oligonucleotide comprises 2, 3, 4, 5, 6 or 7 LNA units. In some aspects, every LNA unit in the oligonucleotide is a stereoisomer in the same configuration. In some aspects, every LNA unit in the oligonucleotide is a beta-D-oxy LNA unit or every LNA unit in the oligonucleotide is an alpha- L-oxy-LNA unit. In some aspects, the sequence of the oligonucleotide comprises at least one phosphorothioate, phosphorodithioate, or boranophosphate internucleoside linkage. In some aspects, one or more of the internucleoside linkages comprises a chiral center in the R conformation and/or in the S conformation. In some aspects, the oligonucleotide comprising an LNA can form a duplex with a human synaptogyrin-3 target sequence selected from the group consisting of SEQ. ID No. 2-93 of with increased thermal stability with respect to a corresponding duplex comprising the corresponding oligonucleotide without LNA. In some aspects, the oligonucleotide of the present disclosure is an RNAi molecule conjugate comprising an RNAi molecule covalently attached to non-nucleotide or non-polynucleotide moiety, which can be attached to the 5' end, 3' end, or both. In some aspects, the non-nucleotide or non-polynucleotide moiety is a targeting moiety that is attached to the 5' -end or to the 3' -end of the RNAi molecule. In some aspects, the targeting moiety is linked to the RNAi molecule via a linker. In some aspects, the targeting moiety comprises a carbohydrate conjugate moiety comprising a carbohydrate selected from the group consisting of galactose, lactose, N-acetylgalactosamine (GalNAc), mannose, mannose-6-phosphate, and combinations thereof. In some aspects, the carbohydrate conjugate moiety is not a linear carbohydrate polymer. In some aspects, the carbohydrate conjugate moiety is a carbohydrate group comprising 1, 2, 3, or 4 carbohydrate moieties. In some aspects, the carbohydrate moieties are identical or non-identical. In some aspects, the carbohydrate conjugate moiety comprises at least one asialoglycoprotein receptor targeting conjugate moiety. In some aspects, the asialoglycoprotein receptor targeting conjugate moiety comprises a monovalent, divalent, trivalent, or tetravalent GalNAc cluster. In some aspects, each GalNAc in the GalNAc cluster is attached to a branch point group via a spacer. In some aspects, the branch point group comprises di-lysine. In some aspects, the spacer comprises a PEG spacer. In some aspects, the linker comprises a C6 to C12 amino alkyl group or a biocleavable phosphate nucleotide linker comprising between 1 to 6 nucleotides. In some aspects, the targeting moiety targets the oligonucleotide of the present disclosure to the central nervous system (CNS). In some aspects, the targeting moiety allow the oligonucleotide of the present disclosure to permeate through the blood-brain-barrier (BBB).

In some aspects, the oligonucleotide of the application, more particularly the RNAi molecule of the application is a double stranded nucleic acid. In some aspects, the RNAi molecule is a siRNA. In some aspects, the RNAi molecule of the present disclosure is a di-siRNA. In some aspects, the RNAi molecule of the present disclosure is a shRNA. In some aspects, the antisense oligomer portion of an oligonucleotide of the present disclosure is an antisense oligonucleotide (ASO). In some aspects, the antisense oligomer portion of an oligonucleotide of the present disclosure is multimeric. In some aspects, the antisense oligomer portion of an oligonucleotide of the present disclosure is a multimeric ASO, e.g., it can comprise several concatenated antisense oligomers of the present disclosure. In some aspects, the antisense oligomer portion of an oligonucleotide of the present disclosure comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more than 10 concatenated antisense oligomers. In some aspects, the concatenated oligomers are connected via cleavable linkers interposed between each ASO unit in the ASO multimer. In some aspects, the antisense oligomer portion of an oligonucleotide of the present disclosure can target a target region in the synaptogyrin-3 mRNA selected from the group consisting of SEQ ID No. 2- 93.

In some aspects, the antisense oligomer portion of an oligonucleotide of the present disclosure comprises a complementarity region that is complementary to at least 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16 nucleotides of a target region in the synaptogyrin-3 mRNA selected from the group consisting of SEQ. ID No. 2-93.

The oligonucleotides of the present disclosure are capable of modulating the expression of the synaptogyrin-3 gene by specifically targeting a targeting region in a synaptogyrin-3 RNA, e.g., a mRNA. In some aspects, the oligonucleotide of the present disclosure is capable of down-regulating expression of the synaptogyrin-3 gene by binding to such target region. Thus, in some aspects, the oligonucleotide of the present disclosure can affect (reduce or inhibit) the expression of synaptogyrin-3, e.g., in a mammalian subject such a human, by binding to a specific target region in a synaptogyrin-3 RNA, e.g., an mRNA. In some aspects, the oligonucleotide of the present disclosure can affect the expression of synaptogyrin-3 in a human cell, by binding to a specific target region in a synaptogyrin-3 RNA, e.g., an mRNA. In some aspects, the RNA is an mRNA, such as pre-mRNA. In some aspects, the RNA is a mature mRNA. The oligonucleotide according to the present disclosure is preferably capable of hybridizing to the target nucleic acid.

In some aspects, the target sequence can extend 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides beyond the 5' end of a synaptorgyrin-3 target region comprising or consisting of a sequence set forth in SEQ ID No. 2- 93. In some aspects, the target sequence can extend 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides beyond the 3' end of a synaptorgyrin-3 target region comprising or consisting of a sequence set forth in SEQ ID No. 2-93. In some aspects, the target sequence can extend 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides beyond the 5' end and/or 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides beyond the 3' end of a synaptogyrin-3 target region comprising or consisting of a sequence set forth in SEQ ID No. 2-93. In some aspects, the extended target region overlaps with 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or 16 nucleotides of a synaptogyrin-3 target region comprising or consisting of a sequences set forth in SEQ ID No. 2-93.

In some aspects, the nucleotides extending beyond the 5' end and/or the 3' end of a sequence set forth in SEQ ID No. 2-93 is complementary (partially or fully complementary) to a corresponding sequence in the mRNA transcript of SEQ ID No. 1. The present disclosure also provides antisense oligonucleotides that are complementary, e.g., fully complementary, to these target sequences. In some aspects, the present disclosure provides a target sequence comprising a 21-mer sequence selected from SEQ ID No. 2-4, 6-15, 17, 18, 20, 21, 23-25, 27-29, 31-41, 43-45, 47-49, 51-57, 60-68, 70- 81, and 83-92, or a 12, 13, 14, 15, 16, 17, 18, 19, or 20-mer subsequence thereof. Furthermore, the present disclosure provides a target sequence comprising (i) a sequence selected from the group consisting of SEQ. ID No. 2-4, 6-15, 17, 18, 20, 21, 23-25, 27-29, 31-41, 43-45, 47-49, 51-57, 60-68, 70-81, and 83-92, and 12-mer, 13-mer, 14-mer, 15-mer, 16-mer, 17-mer, 18-mer, 19-mer, or 20-mer subsequences thereof, plus (ii) 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 additional 5' nucleotides and/or 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 additional 3' nucleotides. In some aspects, the additional 5' and/or 3' nucleotides are complementary (partially or fully complementary) to a corresponding sequence in the mRNA transcript of SEQ ID No. 1. The present disclosure also provides antisense oligonucleotides that are complementary, e.g., fully complementary, to these target sequences.

In some aspects, the target region comprises or consists of a corresponding target sequence region derived from the sequence of a mutant or allelic variant of a human synaptogyrin-3 gene encoding the mRNA transcript of SEQ ID No. 1. In other aspects, the target region can be a subsequence present in another synaptogyrin-3 mRNA transcript variant encoding human synaptogyrin-3. In some aspects, the target region comprises or consists of a corresponding target sequence region derived from the sequence of a paralog or ortholog of the human synaptogyrin-3 gene encoding the mRNA of SEQ ID No. 1.

In some aspects, the target region is within an exon. In some aspects, the target region comprises the junction between and intron and an exon.

In some aspects, the oligonucleotides of the present disclosure bind to the target nucleic acid (e.g., a subsequence of an mRNA transcript wherein the subsequence is selected from the group consisting of SEQ ID No. 2-93 and the effect on synaptogyrin-3 expression and/or activity level is at least about 10% to about 20% reduction in synaptogyrin-3 expression and/or activity level compared to the normal or control synaptogyrin-3 expression level (e.g., the synaptogyrin-3 expression level of a cell, animal or human treated with saline) and/or normal or control synaptogyrin-3 activity level (e.g. the expression level of a cell, animal or human treated with saline). In some aspects, the reduction in synaptogyrin-3 expression and/or activity is at least about 10%, about least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% compared to the normal or control expression and/or activity level. In some aspects, the reduction in synaptogyrin-3 expression and/or activity is about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% compared to the normal or control synaptogyrin-3 expression and/or activity level.

In some aspects, the synaptogyrin-3 expression level and/or protein level and/or activity level after the administration of an oligonucleotide of the present disclosure is less than about 2%, less than about 5%, less than about 10%, less than about 15%, less than about 20%, less than about 25%, less than about 30%, less than about 35%, less than about 40%, less than about 45%, less than about 50%, less than about 55%, less than about 60%, less than about 65%, less than about 70%, less than about 75%, or less than about 80% of the synaptogyrin-3 expression level and/or protein level and/or activity level prior to the administration of an oligonucleotide of the present disclosure.

In some aspects, the synaptogyrin-3 expression level and/or protein level and/or activity level after the administration of an oligonucleotide of the present disclosure is about 2% to about 5%, about 5% to about 10%, about 10% to about 15%, about 15% to about 20%, about 20%, to about 25%, about 25% to about 30%, about 30% to about 35%, about 35% to about 40%, about 40% to about 45%, about 45% to about 50%, about 50% to about 55%, about 55% to about 60%, about 60% to about 65%, about 65% to about 70%, about 70% to about 75%, or about 75% to about 80% of the synaptogyrin-3 expression level and/or protein level and/or activity level prior to the administration of an oligonucleotide of the present disclosure.

The present disclosure therefore provides an in vitro or in vivo method of down-regulating or inhibiting the expression of synaptogyrin-3 protein and/or mRNA transcript in a cell which is expressing synaptogyrin-3 protein and/or mRNA, said method comprising administering an oligonucleotide of the present disclosure, e.g., as a pharmaceutical composition of the present disclosure to said cell to down- regulate or inhibit the expression of synaptogyrin-3 protein and/or mRNA in said cell. Suitably the cell is a mammalian cell such as a human cell.

It is to be understood that in some aspects the oligonucleotides of the present disclosure can be multimers comprising, e.g., 2, 3, 4, 5, 6, or more concatenated oligonucleotides disclosed herein, which can optionally be connected by spacers or linkers comprising nucleotide or non-nucleotide units interposed between each oligonucleotide in the multimer. Accordingly, in some aspects, the oligonucleotides of the present disclosure can comprise or consist of a contiguous nucleotide sequence of a total of at least about 20, at least about 25, at least about 30, at least about 35, at least about 40, at least about 45, at least about 50, at least about 55, at least about 60, at least about 65, at least about 70, at least about 75, at least about 80, at least about 85, at least about 90, at least about 95, at least about 100, at least about 110, at least about 120, at least about 130, at least about 140, at least about 150, at least about 160, at least about 170, at least about 180, at least about 190, or at least about 200 contiguous nucleotides in length.

In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 172, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ. ID No. 173, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 174, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 175, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 176, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 177, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 178, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 179, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 180, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 181, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 182, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 183, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ. ID No. 184, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 185, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 186, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 187, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 188, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 189, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 190, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 191, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 192, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 193, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 194, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 195, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 196, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 197, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 198, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ. ID No. 199, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 200, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 201, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 202, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 203, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 204, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 205, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 206, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 207, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 208, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 209, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 210, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 211, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 212, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ. ID No. 213, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 214, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 215, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 216, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 217, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 218, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 219, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 220, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 221, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 222, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 223, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 224, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 225, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 226, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 227, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ. ID No. 228, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 229, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 230, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 231, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 232, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer.ln some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 233, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 234, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 235, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 236, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 237, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 238, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 239, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 240, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 241, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ. ID No. 242, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 243, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 244, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 245, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 246, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 247, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 248, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects, the oligonucleotide of the present disclosure comprises a single stranded oligomer comprising an antisense sequence set forth in SEQ ID No. 249, or a 12 to 21 contiguous nucleotides subsequence thereof, e.g., a 16-mer. In some aspects of the antisense sequence disclosed above, all odd positions in the antisense sequence comprise a 2'0-Methyl modification and all even positions in the antisense sequence comprise a 2'fluoro modification. In some aspects of the antisense sequences disclosed above the first two 5' and the last two 3' internucleoside linkages are phosphorothioate.

In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 94 and 172. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 95 and 173. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 96 and 174. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 97 and 175. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ. ID No. 98 and 176. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 99 and 177. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 100 and 178. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 101 and 179. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 102 and 180. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 103 and 181. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 104 and 182. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 105 and 183. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 106 and 184. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 107 and 185. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 108 and 186. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 109 and 187. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 110 and 188. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. Ill and 189. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ. ID No. 112 and 190. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 113 and 191. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 114 and 192. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 115 and 193. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 116 and 194. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 117 and 195. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 118 and 196. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 119 and 197. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 120 and 198. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 121 and 199. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 122 and 200. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 123 and 201. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 124 and 202. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ. ID No. 125 and 203. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 126 and 204. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 127 and 205. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 128 and 206. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 129 and 207. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 130 and 208. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 131 and 209. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 132 and 210. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 133 and 211. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 134 and 212. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 135 and 213. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 136 and 214. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 137 and 215. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 138 and 216. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ. ID No. 139 and 217. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 140 and 218. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 141 and 219. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 142 and 220. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 143 and 221. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 144 and 222. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 145 and 223. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 146 and 224. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 147 and 225. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 148 and 226. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 149 and 227. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 150 and 228. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ. ID No. 151 and 229. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 152 and 230. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 153 and 231. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 154 and 232. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 155 and 233. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 156 and 234. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 157 and 235. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 158 and 236. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 159 and 237. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 160 and 238. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 161 and 239. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 162 and 240. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 163 and 241. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 164 and 242. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 165 and 243. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ. ID No. 166 and 244. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 167 and 245. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 168 and 246. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 169 and 247. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 170 and 248. In some aspects, the nucleic acid of the present disclosure is a duplex (e.g., a siRNA or an shRNA) comprising a sense strand and an antisense strand which respectively comprise or consist of the sense and antisense sequences set forth in SEQ ID No. 171 and 249. In some aspects of the nucleic acid duplex sequences disclosed above, the 3' end of the sense strand and the 5' end of the antisense strand are connected by a loop. In some aspects of the nucleic acid duplex sequences disclosed above, all odd positions in the antisense strand sequence comprise a 2'-O-Methyl modification and all even positions in the antisense sequence comprise a 2'-Fluoro modification. In some aspects of the nucleic acid duplex sequences disclosed above, all even positions in the sense sequence comprise a 2'-O-Methyl modification, and all off positions in the sense strand sequence comprise a 2'-Fluoro modification, except the first 5' end nucleotide (position 1) which comprises a 2'-O-Methyl modification. In some aspects of the nucleic acid duplex sequences disclosed above the first two 5' and the last two 3' internucleoside linkages of the sense strand are phosphorothioate. In some aspects of the nucleic acid duplex sequences disclosed above the first two 5' and the last two 3' internucleoside linkages of the antisense strand are phosphorothioate.

The target regions within Synaptogyrin-3 mRNA of the application

It was surprisingly found that some regions within the Synaptogyrin-3 mRNA transcript are significantly more accessible for oligonucleotides such as RNAi molecules and therefore are preferred regions for designing oligonucleotides suitable for or capable of reducing the expression and/or activity of Synaptogyrin-3. By transcript-walking the borders of the identified regions were determined (TABLE 1). The level of Syngr-3 mRNA was determined in SH-SY5Y cells using 20 nM and 2 nM doses of siRNAs comprising a 21-mer antisense sequence and a 19-mer sense sequence. A dose response curve (DRC) was determined for siRNAs that could significantly reduce the Syngr-3 mRNA transcript level.

TABLE 1. Overview of siRNA molecules developed and tested by the inventors of current application listed based on their binding position on the synaptogyrin-3 transcript from 5' to 3'. The symbol "//" represents several siRNA molecules that do not reduce the synaptogyrin-3 transcript level and are not shown due to space limitations.

The threshold for a region suitable for antisense design was set at the level at which a tested siRNA in at least one of the experiments (at a 2 nM or 20 nM dose) would be able to reduce the human Syngr-3 mRNA transcript with at least 15% with respect to a reference system (e.g., baseline levels in an individual or population of individuals, or below a pre-determined threshold value). Nevertheless, therapeutic effects may be obtained at levels of inhibition of the expression of the human Syngr-3 mRNA transcript below 15% as the DRC results consequently showed a stronger reduction of the Syngr-3 transcript level (Table 1). The siRNA molecules selected from the primary screen were retested in two different cell lines (i.e. SH-SY5Y and SKBR3 cells) (TABLE 2). As used herein, the term "reducing", e.g., reducing the level of hSYNGR3 mRNA transcript, of hSYNGR3 protein level or hSYNGR3 activity or a combination thereof, refers to the ability of an oligonucleotide of the present disclosure (e.g., an ASO or siRNA) to statistically significantly reduce (or decrease, inhibit or lower) the level of hSYNGR3 gene transcript (mRNA, e.g. pre-mRNA or mature mRNA) and/or hSYNGR3 protein level and/or activity in a cell, a tissue, or a subject. In some aspects, the term "reducing" refer to complete reduction or inhibition (100% inhibition or non-detectable level) of hSYNGR3 mRNA transcript and/or hSYNGR3 protein level and/or activity. In other aspects, the term "reducing" refers, e.g., to at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least 90%, at least 95% or at least 99% reduction or inhibition of the level of hSYNGRS mRNA transcript and/or hSYNGRS protein expression and/or activity in a cell, a tissue, or a subject.

The terms "individual", "subject", "host", and "patient", are used interchangeably herein and refer to any mammalian subject for whom diagnosis, treatment, or therapy is desired, particularly humans. The compositions and methods described herein are applicable to both human therapy and veterinary applications. In some aspects, the subject is a mammal, and in other aspects the subject is a human. As used herein, a "mammalian subject" includes all mammals, including without limitation, humans, domestic animals (e.g., dogs, cats and the like), farm animals (e.g., cows, sheep, pigs, horses and the like) and laboratory animals (e.g., monkey, rats, mice, rabbits, guinea pigs and the like).

The application thus provides target regions within the human Syngr-3 gene or the Syngr3 mRNA transcript for designing oligonucleotides (e.g. ASO, siRNA or shRNA) that reduce the expression of hSYNGR3 by at least 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least 90%, at least 95% or at least 99% compared to a control situation, e.g. where no oligonucleotide (e.g. ASO, siRNA or shRNA) was administered or where a scrambled control molecule was used as a negative control. Said target regions and the corresponding sequences are shown in FIGURE 3 and 4.

TABLE 2. Confirmation of the IC50 and maximum inhibition of selected siRNA molecules from Table 1 in SH-SY5Y and SKBR3 cells.

In one embodiment, a synaptogyrin-3 mRNA transcript target sequence selected from the group consisting of SEQ. ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 69, 82 and 93 is provided for designing an antisense or RNAi molecule (e.g. ASO, siRNA or shRNA) capable of reducing the level of Synaptogyrin- 3 transcript, the level of expressed synaptogyrin-3, the level of synaptogyrin-3 activity, or a combination thereof in a cell with at least 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least 90%, at least 95% or at least 99% compared to a control situation, e.g., where no oligonucleotide (e.g., ASO, siRNA or shRNA) was administered or where a scrambled control molecule was used as a negative control. In a particular embodiment, said synaptogyrin-3 mRNA transcript target sequence comprises or consists of a sequence selected from the group consisting of SEQ ID No. 2-4, 6-15, 17-18, 20-21, 23-25, 27-29, 31-41, 43-45, 47-49, 51-57, 60-68, 70-81 and SEQ ID No. 83-92.

In another embodiment, the oligonucleotide of the application (described in detail above) comprises a contiguous nucleotide sequence of at least 10 contiguous nucleotides in length which are partially or fully complementary to a target region of Synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1 or allelic variants thereof. In a particular embodiment, the contiguous nucleotide sequence of at least 10 contiguous nucleotides in length is at least 80%, at least 81%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% complementary to a target region in the synaptogyrin-3 mRNA transcript. In a particular embodiment, the contiguous nucleotide sequence of at least 10 contiguous nucleotides in length is about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, about 100% complementary to a target region in the synaptogyrin-3 mRNA transcript. In another particular embodiment, the contiguous nucleotide sequence of at least 10 contiguous nucleotides in length is complementary to an equal length portion of a target region in the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1 or a variant thereof, e.g. an allelic variant thereof.

The target regions of the present disclosure are defined as subsequences of SEQ. ID No. 1, wherein the subsequences are delineated by a 5' -end position and a 3' -end position. For example, a target region between the nucleobases at positions 2 and 10 of SEQ ID No. 1 consists of consecutive nucleobases 2, 3, 4, 5, 6, 7, 8, 9 and 10 of SEQ ID No. 1 or AGGCGGCAG. It is thus to be understood that any range given in the current disclosure includes the range endpoints. Accordingly, if a target region is located between the nucleotides at positions 2 and 10 of SEQ ID No. 1, both nucleotide 2 and nucleotide 10 of SEQ ID No. 1 are included.

As used throughout the present disclosure, the terms "target regions of SEQ ID No. 1" or "target region in the synaptogyrin-3 mRNA transcript" or "target region" in general, as well as grammatical variants thereof refer to regions or subsequences in a synaptogyrin-3 mRNA transcript that are targeted by the oligonucleotides of the present disclosure. In some aspects, the synaptogyrin-3 mRNA transcript containing the target region is the synaptogyrin-3 mRNA transcript set forth in SEQ ID No. 1. However, in other aspects, the synaptogyrin-3 mRNA transcript containing the target region can be a synaptogyrin- 3 mRNA transcript variant, e.g., an allelic variant, of the synaptogyrin-3 mRNA transcript set forth in SEQ ID No. 1, an isoform thereof, or an ortholog thereof.

In a particular aspect, the target region is a subsequence located between nucleobase positions 205 and 265 of the synaptogyrin-3 mRNA transcript set forth in SEQ ID No. 1, or is comprised within the nucleotide subsequence located between nucleobase positions 205 and 265 of the synaptogyrin-3 mRNA transcript set forth in SEQ ID No. 1. In some aspects, the target region is a subsequence located between nucleobase positions 206 and 265, 207 and 265, 208 and 265, 209 and 265, 210 and 265, 211 and 265, 212 and 265, 213 and 265, 214 and 265, 215 and 265, 216 and 265, 217 and 265, 218 and 265, 219 and 265, 220 and 265, 221 and 265, 222 and 265, 223 and 265, 224 and 265, 225 and 265, 226 and 265, 227 and 265, 228 and 265, 229 and 265, 205 and 258, 205 and 259, 205 and 260, 205 and 261, 205 and 262, 205 and 263, or 205 and 264 of the synaptogyrin-3 mRNA transcript set forth in SEQ ID No. 1. More particularly, the target region is located between nucleobase positions 224 and 263 of the synaptogyrin- 3 mRNA transcript set forth in SEQ ID No. 1, more particularly between nucleobase positions 225 and 262, 226 and 261, 227 and 260 or 228 and 259, even more particularly between nucleobase positions 229 and 258 of the synaptogyrin-3 mRNA transcript set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 5. In an even more particular embodiment, the target region is defined by (or comprises of consists of) SEQ ID No. 2, 3 and/or 4. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 94, 95, 96, 172, 173 and/or SEQ. ID No. 174.

In another particular embodiment, the target region is located between nucleobase positions 255 and 348 or is comprised within the nucleotide subsequence defined by nucleobase positions 255 and 348 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 256 and 348, 257 and 348, 258 and 348, 259 and 348, 260 and 348, 261 and 348, 262 and 348, 263 and 348, 264 and 348, 265 and 348, 255 and 325, 255 and 326, 255 and 327, 255 and 328, 255 and 329, 255 and 330, 255 and 331, 255 and 332, 255 and 333, 255 and 334, 255 and 335, 255 and 336, 255 and 337, 255 and 338, 255 and 339, 255 and 340, 255 and 341, 255 and 342, 255 and 343, 255 and 344, 255 and 345, 255 and 346, or 255 and 347 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly between nucleobase positions 260 and 330 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 261 and 329, 262 and 328, 263 and 327 or 264 and 326, even more particularly between nucleobase positions 265 and 325 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 16. In an even more particular embodiment, the target region is defined by (or comprises or consists of) a sequence selected from the group consisting of SEQ ID No. 6, 7, 8, 9, 10, 11, 12, 13, 14 and 15. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 175, 176, 177, 178, 179, 180, 181, 182, 183 and/or SEQ ID No. 184. In another particular embodiment, the target region is located between nucleobase positions 338 and 387 or is comprised within the nucleotide subsequence defined by nucleobase positions 338 and 387 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 339 and 387, 340 and 387, 341 and 387, 342 and 387, 343 and 387, 338 and 377, 338 and 378, 338 and 379, 338 and 380, 338 and 381, 338 and 382, 338 and 383, 338 and 384, 338 and 385, or 338 and 386 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly between nucleobase positions 338 and 382 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 339 and 381, 340 and 380, 341 and 379, 342 and 378, even more particularly between nucleobase positions 343 and 377 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 19. In an even more particular embodiment, the target region is defined by (or comprises or consists of) a sequence selected from the group consisting of SEQ ID No. 17 and 18. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 107, 108, 185 and/or SEQ ID No. 186. In another particular embodiment, the target region is located between nucleobase positions 369 and 433 or is comprised within the nucleotide subsequence defined by nucleobase positions 369 and 433 of the synaptogyrin-3 mRNA transcript set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobases 370 and 433, 371 and 433, 372 and 433, 373 and 433, 374 and 433, 375 and 433, 376 and 433, 377 and 433, 378 and 433, 379 and 433, 380 and 433, 381 and 433, 382 and 433, 383 and 433, 384 and 433, 385 and 433, 386 and 433, 387 and 433, 388 and 433, 389 and 433, 390 and 433, 391 and 433, 392 and 433, 393 and 433, 369 and 427, 369 and 428, 369 and 429, 369 and 430, 369 and 431, or 369 and 432 of the synaptogyrin-3 mRNA transcript as set forth in SEQ. ID No. 1. More particularly that target region is located between nucleobase positions 388 and 432 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 389 and 431, 390 and 430, 391 and 429, 392 and 428, even more particularly the target region is located between nucleobase positions 393 and 427 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 22. In an even more particular embodiment, the target region is defined by (or comprises or consists of) SEQ ID No. 20 and/or 21. In a most particular embodiment, the oligonucleotide comprises or consists of SEQ ID No. 109, 110, 187 and/or SEQ ID No. 188.

In another particular embodiment, the target region is located between nucleobase positions 422 and 531 or is comprised within the nucleotide subsequence defined by nucleobase positions 422 and 531 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 423 and 531, 424 and 531, 425 and 531, 426 and 531, 427 and 531, 428 and 531, 429 and 531, 430 and 531, 431 and 531, 432 and 531, 433 and 531, 434 and 531, 422 and 526, 422 and 527, 422 and 528, 422 and 529, or 422 and 530 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly the target region is located between nucleobase positions 429 and 531 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 430 and 530, 431 and 529, 432 and 528, 433 and 527, even more particularly the target regions is located between nucleobase positions 434 and 526 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 26. In an even more particular embodiment, the target region is defined by (or comprises or consists of) SEQ ID No. 23, 24 and/or 25. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. Ill, 112, 113, 189, 190 and/or SEQ ID No. 191. In another particular embodiment, the target region is located between nucleobase positions 603 and 656 or is comprised within the nucleotide subsequence defined by nucleobase positions 603 and 656 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 604 and 656, 605 and 656, 606 and 656, 607 and 656, 608 and 656, 609 and 656, 610 and 656, 603 and 632, 603 and 633, 603 and 634, 603 and 635, 603 and 636, 603 and 637, 603 and 638, 603 and 639, 603 and 640, 603 and 641, 603 and 642, 603 and 643, 603 and 644, 603 and 645, 603 and 646, 603 and 647, 603 and 648, 603 and 649, 603 and 650, 603 and 651, 603 and 652, 603 and 653, 603 and 654, or 603 and 655 of the synaptogyrin-3 mRNA transcript as set forth in SEQ. ID No. 1. More particularly the target region is located between nucleobase positions 605 and 637 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 606 and 636, 607 and 635, 608 and 634, 609 and 633, even more particularly the target region is located between nucleobase positions 610 and 632 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 30. In an even more particular embodiment, the target region is defined by (or comprises or consists of) SEQ ID No. 27, 28 and/or 29. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 114, 115, 116, 192, 193 and/or SEQ ID No. 194.

In another particular embodiment, the target region is located between nucleobase positions 641 and 714 or is comprised within the nucleotide subsequence defined by nucleobase positions 641 and 714 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 642 and 714, 643 and 714, 644 and 714, 645 and 714, 646 and 714, 647 and 714, 648 and 714, 649 and 714, 650 and 714, 641 and 683, 641 and 684, 641 and 685, 641 and 686,

641 and 687, 641 and 688, 641 and 689, 641 and 690, 641 and 691, 641 and 692, 641 and 693, 641 and

694, 641 and 695, 641 and 696, 641 and 697, 641 and 698, 641 and 699, 641 and 700, 641 and 701, 641 and 702, 641 and 703, 641 and 704, 641 and 705, 641 and 706, 641 and 707, 641 and 708, 641 and 709,

641 and 710, 641 and 711, 641 and 712, or 641 and 713 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly the target region is located between nucleobase position 645 and 688 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 646 and 687, 647 and 686, 648 and 685, 649 and 684, even more particularly the target regions is located between nucleobase positions 650 and 683 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 42. In an even more particular embodiment, the target region is defined by (or comprises or consists of) SEQ ID No. 31, 32, 33, 34, 35, 36, 37, 38, 39, 40 and/or 41. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204 and/or SEQ. ID No. 205.

In another particular embodiment, the target region is located between nucleobase positions 717 and 768 or is comprised within the nucleotide subsequence defined by nucleobase positions 717 and 768 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 718 and 768, 719 and 768, 720 and 768, 721 and 768, 722 and 768, 723 and 768, 724 and 768, 725 and 768, 726 and 768, 1 1 and 768, 728 and 768, 729 and 768, 730 and 768,

731 and 768, 732 and 768, 733 and 768, 734 and 768, 735 and 768, 736 and 768, 737 and 768, 717 and 762, 717 and 763, 717 and 764, 717 and 765, 717 and 766, or 717 and 767 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly the target region is located between nucleobase

732 and 767 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 733 and 766, 734 and 765, 735 and 764, 736 and 763, even more particularly the target region is located between nucleobase 737 and 762 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 46. In an even more particular embodiment, the target region is defined by (or comprises of consists of) SEQ ID No. 43, 44 and/or 45. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 128, 129, 130, 206, 207 and/or SEQ ID No. 208.

In another particular embodiment, the target region is located between nucleobase positions 1150 and 1600 or is comprised within the nucleotide subsequence defined by nucleobase positions 1150 and 1600 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 1151 and 1600, 1152 and 1600, 1153 and 1600, 1154 and 1600, 1155 and 1600, 1156 and 1600, 1157 and 1600, 1158 and 1600, 1159 and 1600, 1160 and 1600, 1161 and 1600, 1162 and 1600, 1163 and 1600, 1164 and 1600, 1165 and 1600, 1166 and 1600, 1167 and 1600, 1168 and 1600, 1169 and 1600, 1170 and 1600, 1171 and 1600, 1172 and 1600, 1173 and 1600, 1174 and 1600, 1175 and 1600, 1176 and 1600, 1177 and 1600, 1178 and 1600, 1179 and 1600, 1180 and 1600, 1181 and 1600, 1182 and 1600, 1183 and 1600, 1183 and 1600, 1184 and 1600, 1185 and 1600, 1186 and 1600, 1187 and 1600, 1188 and 1600, 1189 and 1600, 1190 and 1600, 1191 and 1600, 1192 and 1600, 1193 and 1600, 1194 and 1600, 1195 and 1600, 1196 and 1600, 1197 and 1600, 1198 and 1600, 1199 and 1600, 1200 and 1600, 1201 and 1600, 1202 and 1600, 1203 and 1600, 1204 and 1600, 1205 and 1600, 1206 and 1600, 1207 and 1600, 1208 and 1600, 1209 and 1600, 1210 and 1600, 1211 and 1600, 1212 and 1600, 1213 and 1600, 1214 and 1600, 1215 and 1600, 1216 and 1600, 1217 and 1600, 1218 and 1600, 1219 and 1600, 1220 and 1600, 1221 and 1600, 1222 and 1600, 1223 and 1600, 1224 and 1600, 1225 and 1600, 1226 and 1600, 1227 and 1600, 1228 and 1600, 1229 and 1600, 1230 and 1600, 1231 and 1600, 1232 and 1600, 1233 and 1600, 1234 and 1600, 1235 and 1600, 1236 and 1600, 1237 and 1600, 1238 and 1600, 1239 and 1600, 1240 and 1600, 1241 and 1600, 1242 and 1600, 1243 and 1600, 1244 and 1600, 1245 and 1600, 1246 and 1600, 1247 and 1600, 1248 and 1600, 1249 and 1600, 1250 and 1600, 1251 and 1600, 1252 and 1600, 1253 and 1600, 1254 and 1600, 1255 and 1600, 1256 and 1600, 1257 and 1600, 1258 and 1600, 1259 and 1600, 1260 and 1600, 1261 and 1600, 1262 and 1600, 1263 and 1600, 1264 and 1600, 1265 and 1600, 1266 and 1600, 1267 and 1600, 1150 and 1535, 1150 and 1536, 1150 and 1537, 1150 and 1538, 1150 and 1539, 1150 and 1540, 1150 and 1541, 1150 and 1542, 1150 and 1543, 1150 and 1544, 1150 and 1545, 1150 and 1546, 1150 and 1547, 1150 and 1548, 1150 and 1549, 1150 and 1550, 1150 and 1551, 1150 and 1552, 1150 and 1553, 1150 and 1554, 1150 and 1555, 1150 and 1556, 1150 and 1557, 1150 and 1558, 1150 and 1559, 1150 and 1560, 1150 and 1561, 1150 and 1562, 1150 and 1563, 1150 and 1564, 1150 and 1565, 1150 and 1567, 1150 and 1568, 1150 and 1569, 1150 and 1570, 1150 and 1571, 1150 and 1572, 1150 and 1573, 1150 and 1574, 1150 and 1575, 1150 and 1576, 1150 and 1577, 1150 and 1578, 1150 and 1579, 1150 and 1580, 1150 and 1581, 1150 and 1582, 1150 and 1583, 1150 and 1584, 1150 and 1585, 1150 and 1586, 1150 and 1587, 1150 and 1588, 1150 and 1589, 1150 and 1590, 1150 and 1591, 1150 and 1592, 1150 and 1593, 1150 and 1594, 1150 and 1595, 1150 and 1596, 1150 and 1597, 1150 and 1598, or 1150 and 1599 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly the target region is located between nucleobase positions 1262 and 1540 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 1263 and 1539, 1264 and 1538, 1265 and 1537, or 1266 and 1536, even more particularly the target region is located between nucleobase positions 1267 and 1535 of the synaptogyrin-3 mRNA transcript as set forth in SEQ. ID No. 1. In a more particular embodiment, the target region is defined by or set forth in SEQ ID No. 59.

In another particular embodiment, the target region is located between nucleobase 1262 and 1307 or is comprised within the nucleotide subsequence defined by nucleobase positions 1262 and 1307 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 1263 and 1307, 1264 and 1307, 1265 and 1307, 1266 and 1307, 1267 and 1307, 1262 and 1302, 1262 and 1303, 1262 and 1304, 1262 and 1305, or 1262 and 1306 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly the target region is located between nucleobase positions 1267 and 1302 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. In a more particular embodiment, the target region is defined by or is set forth in SEQ ID No. 50. In an even more particular embodiment, the target region is defined by (or comprises of consists of) SEQ. ID No. 47, 48 and/or 49. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 131, 132, 133, 209, 210 and/or SEQ ID No. 211.

In another particular embodiment, the target region is located between nucleobase positions 1384 and 1540 or is comprised within the nucleotide subsequence defined by nucleobase positions 1384 and 1540 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobases 1385 and 1540, 1386 and 1540, 1387 and 1540, 1388 and 1540, 1389 and 1540, 1384 and 1535, 1384 and 1536, 1384 and 1537, 1384 and 1538, or 1384 and 1539 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly the target region is located between nucleobase positions 1389 and 1535 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 58. In an even more particular embodiment, the target region is defined by (or comprises or consists of) SEQ ID No. 51, 52, 53, 54, 55, 56 and/or 57. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 134, 135, 136, 137, 138, 139, 140, 212, 213, 214, 215, 216, 217 and/or SEQ ID No. 218.

In another particular embodiment, the target region is located between nucleobase positions 1600 and 2026 or is comprised within the nucleotide subsequence defined by nucleobase positions 1600 and 2026 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 1601 and 2026, 1602 and 2026, 1603 and 2026, 1604 and 2026, 1605 and 2026, 1606 and 2026, 1607 and 2026, 1608 and 2026, 1609 and 2026, 1610 and 2026, 1611 and 2026, 1612 and 2026, 1613 and 2026, 1614 and 2026, 1615 and 2026, 1616 and 2026, 1617 and 2026, 1618 and 2026, 1619 and 2026, 1620 and 2026, 1621 and 2026, 1622 and 2026, 1623 and 2026, 1624 and 2026, 1625 and 2026, 1626 and 2026, 1627 and 2026, 1628 and 2026, 1629 and 2026, 1630 and 2026, 1631 and 2026, 1632 and 2026, 1633 and 2026, 1634 and 2026, 1635 and 2026, 1636 and 2026, 1637 and 2026, 1638 and 2026, 1639 and 2026, 1640 and 2026, 1641 and 2026, 1642 and 2026, 1643 and 2026, 1644 and 2026, 1645 and 2026, 1646 and 2026, 1647 and 2026, 1648 and 2026, 1649 and 2026, 1650 and 2026, 1651 and 2026, 1652 and 2026, 1653 and 2026, 1654 and 2026, 1655 and 2026, 1656 and 2026, 1657 and 2026, 1658 and 2026, 1659 and 2026, 1660 and 2026, 1661 and 2026, 1662 and 2026, 1663 and 2026, 1664 and 2026, 1665 and 2026, 1666 and 2026, 1667 and 2026, 1668 and 2026, 1669 and 2026, 1670 and 2026, 1671 and 2026, 1672 and 2026, 1673 and 2026, 1674 and 2026, 1675 and 2026, 1676 and 2026, 1677 and 2026, 1678 and 2026, 1679 and 2026, 1680 and 2026, 1681 and 2026, 1682 and 2026, 1683 and 2026, 1684 and 2026, 1685 and 2026, 1686 and 2026, 1687 and 2026, 1688 and 2026, 1689 and 2026, 1690 and 2026, 1691 and 2026, 1692 and 2026, 1693 and 2026, 1694 and 2026, 1695 and 2026, 1696 and 2026, 1697 and 2026, 1698 and 2026, 1699 and 2026, 1700 and 2026, 1701 and 2026, 1702 and 2026, 1703 and 2026, 1704 and 2026, 1705 and 2026, 1706 and 2026, 1707 and 2026, 1708 and 2026, 1709 and 2026, 1710 and 2026, 1711 and 2026, 1712 and 2026, 1713 and 2026, or 1600 and 2025 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly the target region is located between nucleobase positions 1708 and 2026 of the synaptogyrin-3 mRNA transcript as set forth in SEQ. ID No. 1, even more particularly between nucleobase positions 1713 and 2025 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 69. In an even more particular embodiment, the target region is defined by (or comprises or consists of) SEQ ID No. 60, 61, 62, 63, 64, 65, 66, 67 and/or 68. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 141, 142, 143, 144, 145, 146, 147, 148, 149, 219, 220, 221, 222, 223, 224, 225, 226 and/or SEQ ID No. 227.

In another particular embodiment, the target region is located between nucleobase positions 1743 and 1868 or is comprised within the nucleotide subsequence defined by nucleobase positions 1743 and 1868 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 1744 and 1868, 1745 and 1868, 1746 and 1868, 1747 and 1868, 1748 and 1868, 1749 and 1868, 1750 and 1868, 1751 and 1868, 1752 and 1868, 1753 and 1868, 1754 and 1868, 1755 and 1868, 1756 and 1868, 1757 and 1868, 1758 and 1868, 1759 and 1868, 1760 and 1868, 1761 and 1868, 1762 and 1868, 1763 and 1868, 1764 and 1868, 1765 and 1868, 1766 and 1868, 1767 and 1868, 1768 and 1868, 1769 and 1868, 1743 and 1865, 1743 and 1866, or 1743 and 1867 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. More particularly the target region is located between nucleobase position 1764 and 1868 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 1765 and 1868, 1766 and 1868 or 1767 and 1867, 1768 and 1866, even more particularly the target region is located between nucleobase positions 1769 and 1865 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 82. In an even more particular embodiment, the target region is defined by (or comprises or consists of) SEQ ID No. 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80 and/or 81. In a most particular embodiment, the oligonucleotide (e.g. ASO, siRNA, shRNA) comprises or consists of SEQ ID No. 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238 and/or SEQ ID No. 239. In another particular embodiment, the target region is located between nucleobase positions 1865 and 2026 or is comprised within the nucleotide subsequence defined by nucleobase positions 1865 and 2026 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 1866 and 2026, 1867 and 2026, 1868 and 2026, 1869 and 2026, 1870 and 2026, 1871 and 2026, 1872 and 2026, 1873 and 2026, 1874 and 2026, 1875 and 2026, 1876 and 2026, 1877 and 2026, 1878 and 2026, 1879 and 2026, 1880 and 2026, 1881 and 2026, 1882 and 2026, 1883 and 2026, 1884 and 2026, 1885 and 2026, 1886 and 2026, 1887 and 2026, 1888 and 2026, 1889 and 2026, 1890 and 2026, 1891 and 2026, 1892 and 2026, 1893 and 2026, 1894 and 2026, 1895 and 2026, 1896 and 2026, 1897 and 2026, 1898 and 2026, 1899 and 2026, 1900 and 2026, 1901 and 2026, 1902 and 2026, 1903 and 2026, 1904 and 2026, 1905 and 2026, 1906 and 2026, 1907 and 2026, 1908 and 2026, 1909 and 2026, 1910 and 2026, 1911 and 2026, 1912 and 2026, 1913 and 2026, 1914 and 2026, 1915 and 2026, 1916 and 2026, 1917 and 2026, 1918 and 2026, 1919 and 2026, 1920 and 2026, 1921 and 2026, 1922 and 2026, 1923 and 2026, 1924 and 2026, 1925 and 2026, 1926 and 2026, 1927 and 2026, 1928 and 2026, 1929 and 2026, 1930 and 2026, 1931 and 2026, 1932 and 2026, 1933 and 2026, 1934 and 2026, 1935 and 2026, 1936 and 2026, 1937 and 2026, 1938 and 2026, 1939 and 2026, 1940 and 2026, or 1865 and 2025 of the synaptogyrin-3 mRNA transcript as set forth in SEQ. ID No. 1. More particularly the target region is located between nucleobase positions 1935 and 2026 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1, more particularly between nucleobase positions 1936 and 2026, 1937 and 2026, 1938 and 2026 or 1939 and 2026 even more particularly between nucleobase positions 1940 and 2025 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1.

In a more particular embodiment, the target region is defined by SEQ ID No. 93. In an even more particular embodiment, the target region is defined by (or comprises of consists of) SEQ ID No. 83, 84, 85, 86, 87, 88, 89, 90, 91 and/or 92. In a most particular embodiment, the oligonucleotide (e.g. siRNA, ASO, shRNA) comprises or consists of SEQ ID No. 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 240, 241, 242, 243, 244, 245, 246, 247, 248 and/or SEQ ID No. 249.

In another particular embodiment, the target region is located between nucleobase positions 1992 and 2026 or is comprised within the nucleotide subsequence defined by nucleobase positions 1992 and 2026 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. Particularly, the target region is located between nucleobase positions 1993 and 2026, 1994 and 2026, 1995 and 2026, 1996 and 2026, 1997 and 2026, or 1992 and 2025 of the synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1. In a more particular embodiment, the target region is defined by (or comprises of consists of) SEQ ID No. 90, 91 and/or 92. In a most particular embodiment, the oligonucleotide (e.g. siRNA, ASO, shRNA) comprises or consists of SEQ. ID No. 169, 170, 171, 247, 248 and/or SEQ ID No. 249.

In some aspects, the present disclosure provides a target region within the human Syngr-3 gene, particularly within the Synaptogyrin-3 mRNA transcript as depicted in SEQ ID No. 1, that can be used for designing oligonucleotides or RNAi molecules (e.g. ASO, siRNA, shRNA) that are capable of reducing the expression level of Synaptogyrin-3 mRNA transcript in a cell, wherein the target region is any of the above described target regions.

Also any of the target regions as described herein, for example a target region set forth in any of SEQ ID No. 2-93, a subsequence thereof, a subsequence of synaptogyrin-3 mRNA transcript as set forth in SEQ ID No. 1 comprising or overlapping with a target region set forth in any of SEQ ID No. 2-93 is provided for designing antisense molecules or RNAi molecules, more particularly siRNA, di-siRNA or shRNA duplexes, that can statistically significantly reduce the expression level of the Synaptogyrin-3 mRNA transcript as depicted in SEQ ID No. 1, the synaptogyrin-3 protein level, the synaptogyrin-3 activity level, or any combination thereof in a cell.

Antisense Oligonucleotides (ASO): In some aspects, the oligonucleotide of the present disclosure is an ASO. Thus, in some aspects, an oligonucleotide of the present disclosure comprises an antisense oligonucleotide (ASO), e.g., an unconjugated or conjugated ASO. Antisense oligonucleotides or ASOs are small (generally, between about 16 nucleotides and about 30 nucleotides or shorter, e.g., between about 12 and about 20 nucleotides), synthetic, single-stranded nucleic acid polymers of diverse chemistries, which can be employed to modulate gene expression via various mechanisms. ASOs can be subdivided into two major categories: RNase H competent and steric block ASOs. In some aspects, the oligonucleotide of the present disclosure is RNase H competent. In some aspects, the oligonucleotide of the present disclosure is a steric block ASO. In some aspects, the oligonucleotide of the present disclosure is a gapmer. Gapmer designs are disclosed, e.g., in WO 2007/146511A2, which is herein incorporated by reference in its entirety.

ASOs can also modulate gene expression by steric hindrance or occupancy and only mechanisms. Steric block oligonucleotides are designed to bind to target transcripts with high affinity but do not induce target transcript degradation as they lack RNase H competence. Such oligonucleotides therefore comprise either nucleotides that do not form RNase H substrates when paired with RNA or a mixture of nucleotide chemistries such that runs of consecutive DNA-like bases are avoided. Steric block oligonucleotides can mask specific sequences within a target transcript and thereby interfere with transcript RNA-RNA and/or RNA-protein interactions. The most widely used application of steric block ASOs is in the modulation of alternative splicing in order to selectively exclude or retain a specific exon(s) in order to disrupt the translation of the target gene. ASOs can also be designed to interfere with maturation and stability of the RNA transcript or to block its interaction with the translation apparatus. In case the ASO can enter the nucleus, mRNA maturation can be modulated by inhibition of 5'-cap formation, inhibition of mRNA splicing or activation of RNaseH (Chan et al 2006 Clin Exp Pharmacol Physiol 33:533-540; this reference also describes some of the software available for assisting in design of ASOs).

In some aspects, the ASO comprises an antisense oligomer 16 to 22 contiguous nucleotides in length, wherein the sequence of the antisense oligomer comprises a contiguous sequence 16 to 22 (e.g., 16) nucleotides in length which is 100% complementary to a sequence or to a subsequence of synaptogyrin- 3 target sequence selected from the group consisting of SEQ ID No. 2-4, 6-15, 17, 18, 20, 21, 23-25, TITS, 31-41, 43-45, 47-49, 51-57, 60-68, 70-81, and 83-92 or from the group consisting of SEQ. ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 69, 82, and 93, wherein the antisense oligomer targets an RNA encoding synaptogyrin-3.

In some aspects, the ASO comprises an antisense oligomer 10 to 16 contiguous nucleotides in length, wherein the sequence of the antisense oligomer comprises a contiguous sequence 10 to 25 nucleotides in length which is 100% complementary to a sequence or to a subsequence of synaptogyrin-3 target sequence selected from the group consisting of SEQ ID No. 2-4, 6-15, 17, 18, 20, 21, 23-25, 27-29, 31-41, 43-45, 47-49, 51-57, 60-68, 70-81, and 83-92, or from the group consisting of SEQ ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 69, 82, and 93, wherein the antisense oligomer targets an RNA encoding synaptogyrin-3.

In some aspects, the ASO is a gapmer. In some aspects, the ASO is conjugated to a targeting moiety, e.g., to a GalNAc moiety.

RNAi using duplex silencers: In some aspects, the oligonucleotide of the present disclosure is an RNAi molecule, particularly an RNAi duplex or a double stranded RNAi oligonucleotide, more particularly the antisense portion or the antisense strand of an RNAi duplex (double stranded RNA). RNA interference (RNAi) is a mechanism by which double-stranded RNA triggers the loss of a homologous RNA molecule. Short interfering RNA (siRNA) molecules are the effector molecules of RNAi and classically consist of an RNA duplex (or alternatively phrased a duplex of RNA molecules) with a length of 21 nucleotides, i.e. 19 complementary bases and 2 terminal 3' overhangs. One of the strands of the siRNA (the guide or antisense strand) is complementary to a target transcript, whereas the other strand is designated the passenger or sense strand. siRNAs act to guide the Argonaute2 protein (AG02), as part of the RNA- induced silencing complex (RISC), to complementary target transcripts. Complete complementarity between the siRNA and the target transcript results in cleavage of the target opposite position of the guide strand, catalysed by AG02, leading to gene silencing.

In a siRNA, the sense strand meets the formal definition of a drug delivery device: it is non-covalently bound, enhances the stability of the antisense strand and must be removed by the Ago2 loading complex before the pharmacophore, the antisense strand, is active.

Numerous variations of the archetypal siRNA design have been developed in terms of reduced passenger strand activity and/or improved potency. These include Dicer substrate siRNAs, small internally segmented siRNAs, self-delivering siRNAs (asymmetric and hydrophobic), single-stranded siRNAs and divalent siRNAs.

In some aspects, the oligonucleotide of the present disclosure is the antisense portion of a shRNA. Short hairpin RNAs (shRNAs) are artificial RNA molecules that are transcribed as a single stranded RNA but because of internal complementarity form a loop or hairpin-like structure. The hairpin is subsequently processed to an siRNA and also leads to the degradation of mRNAs in a sequence-specific manner dependent upon complementary binding of the target mRNA. shRNAs are slightly larger than siRNA molecules and, unlike siRNAs, are produced inside the cell in the nucleus.

Other non-limiting examples of RNAi-mediated duplex silencers are miRNAs and di-siRNAs. microRNAs (miRNAs) are endogenous non-coding RNA molecules that trigger RNAi and that have been implicated in a multitude of physiological and pathophysiological processes. miRNA hairpins embedded within long primary miRNA transcripts are sequentially processed by two RNase III family enzymes, DICER1 (Dicer) and DROSHA, which liberate the hairpin and then cleave the loop sequence, respectively. The resulting duplex RNA is analogous to an siRNA and is then loaded into an Argonaute protein (for example, AG02) while one strand is discarded to generate the mature, single-stranded miRNA species. As with siRNAs, miRNAs guide RISC to target sequences where they initiate gene silencing. In contrast with siRNAs, miRNAs typically bind with partial complementarity and induce silencing via slicer-independent mechanisms.

In some aspects, the oligonucleotide of the present disclosure is a di-siRNA. Divalent siRNAs (di-siRNAs) are recently developed RNA silencing agent alternatives and have been shown to support a potent, sustained gene silencing in the central nervous system of mice and non-human primates following a single injection into the cerebrospinal fluid (Alterman et al 2019 Nature Biotech 37, 884-894). Di-siRNAs are composed of two fully chemically modified, phosphorothioate-containing siRNAs connected by a linker. In some aspects, the siRNA of the present disclosure comprises an antisense strand 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 1 , 28, 29 or 30 contiguous nucleotides in length, wherein the sequence of the antisense strand comprises a contiguous sequence of 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27 , 28, 29 or 30 nucleotides in length which is 100% complementary to a sequence or to a subsequence of synaptogyrin-3 target sequence selected from the group consisting of SEQ ID No. 2-4, 6-15, 17, 18, 20, 21, 23-25, 27-29, 31-41, 43-45, 47-49, 51-57, 60-68, 70-81, and 83-92 or from the group consisting of SEQ. ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 69, 82, and 93, wherein the siRNA targets an RNA encoding synaptogyrin-3.

In some aspects, the siRNA of the present disclosure is conjugated to a targeting moiety, e.g., to a GalNAc moiety.

CRISPR gRNA: Another recent genome editing technology is the CRISPR/Cas system, which can be used to achieve RNA-guided genome engineering. CRISPR interference is a genetic technique which allows for sequence-specific control of gene expression in prokaryotic and eukaryotic cells. It is based on the bacterial immune system-derived CRISPR (clustered regularly interspaced palindromic repeats) pathway. Recently, it was demonstrated that the CRISPR-Cas editing system can also be used to target RNA. It has been shown that the Class 2 type Vl-A CRISPR-Cas effector C2c2 can be programmed to cleave singlestranded RNA targets carrying complementary protospacers (Abudayyeh et al 2016 Science 353/science.aaf5573). C2c2 is a single-effector endoRNase mediating ssRNA cleavage once it has been guided by a single crRNA guide toward the target RNA. Hence, the invention disclosed herein can also be applied to develop gRNAs specifically reducing the expression of Synaptogyrin-3 using the CRISPR/Cas system. Therefore, the application also provides that any of the oligonucleotides herein provided can be used as a gRNA or CRISPR gRNA, more particularly a gRNA or CRISPR gRNA is provided with at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or about 100% complementary to an equal length of a target region of Synaptogyrin-3 as depicted in SEQ ID No. 1, wherein the target region is selected from any of target regions of the application or subsequences thereof.

In some aspects, a gRNA or CRISPR gRNA provided herein has at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% complementary to an equal length of a target region of synaptogyrin-3 as depicted in SEQ ID No. 1, wherein the target region is located between nucleobase position 205 and 265, 255 and 348, 338 and 387, 369 and 433, 422 and 531, 603 and 656, 641 and 714, 717 and 768, 1150 and 1600, 1743 and 1868 or between nucleobase position 1865 and 2026 of SEQ ID No. 1 and wherein the endpoints are included, or a subsequence thereof. In some aspects, a gRNA or CRISPR gRNA provided herein has at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% complementary to an equal length of a target region of synaptogyrin-3 as depicted in SEQ ID No. 1, wherein the target region is selected from any of SEQ. ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 69, 82 or 93, more particularly the target region is selected from any of SEQ ID No. 2-4, 6-15, 17-18, 20-21, 23-25, 27-29, 31-41, 43-45, 47-49, 51-57, 70-81, and 83-92.

In some aspects, the gRNA comprises or consists of a sequence selected from the group consisting of SEQ ID No. 2-4, 6-15, 17-18, 20-21, 23-25, 27-29, 31-41, 43-45, 47-49, 51-57, 70-81, and 83-92. In some aspects, the gRNA comprises or consists of a sequence comprising a subsequence selected from the target regions set forth in SEQ ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 69, 82, and 93.

In a particular embodiment, said gRNA is 10 to 50 or 10 to 40 or 10 to 30 nucleotides in length. For example, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 nucleotides in length.

In another particular embodiment, the gRNA comprises a contiguous nucleotide sequence of at least 10 contiguous nucleotides in length comprising a sequence selected from the group consisting of SEQ ID No. 172-249.

In yet another particular embodiment, the gRNA of the present disclosure comprises a sequence that overlaps with a 9, 10, 11, 12, 13, 14, 15, or 16 nucleobase subsequence from a sequence selected from the group consisting of SEQ ID No. 172-249.

Chemical modifications

In some aspects, the oligonucleotides of the present disclosure comprise non-naturally occurring nucleotide analogues, e.g., nucleotides which have modified sugar moieties, such as bicyclic nucleotides or 2' modified nucleotides, such as 2' substituted nucleotides. An essential step in the evolution of the antisense technology was the creation, innovation and evaluation of the medicinal chemistry of oligonucleotides. The goals were to enhance the affinity for the target sequence (thereby increasing potency), assure effective distribution to peripheral tissues, enhance the duration of action by increasing resistance to degradation by nucleases, improve pharmacokinetic characteristics, reduce the class generic (chemically based) toxicities of the chemical classes widely used for therapeutics, and create designs that support multiple post-binding mechanisms, thereby broadening the utility of the technology.

Within the antisense field a broad effort was initiated to modify essentially every position in a dinucleotide except those required for Watson-Crick base pairing. Thousands of analogues have been synthesized and evaluated so far, and novel analogues continue to be investigated. Three major classes of modifications can be distinguished: modifications of the internucleotide linkage, alterations of the ribose sugar and bioconjugations with for example GalNAc.

Phosphorothioates

In some aspects, the oligonucleotides of the present disclosure comprise one or more non-cleavable internucleotide linkages, e.g., phosphorothioate linkages. The phosphodiester backbone of unmodified DNA and RNA oligonucleotides is highly susceptible to degradation by nucleases in vivo. So, to develop oligonucleotides for therapeutic applications, it was necessary to identify backbone modifications that reduce their susceptibility to nuclease degradation while not compromising other key characteristics such as RNase Hl activation and RNA binding too much.

In phosphorothioate (PS) linkages, a non-bridging oxygen in the phosphate group is substituted by sulfur. The PS moiety provides significant protection against nucleases. Importantly, because of the impact of the greater size of sulfur compared with oxygen, the negative charge of the PS moiety at physiological pH is more widely distributed than in a phosphodiester (PO) moiety. This increases the lipophilicity of oligonucleotides that contain PS moieties, facilitating binding to proteins and thereby preventing rapid excretion of the oligonucleotides by the kidney and facilitating uptake of oligonucleotides into cells and tissues. The PS moiety is the most widely used backbone modification in ASOs and RNAi molecules such as siRNAs.

Ribose sugar modifications

In some aspects, the oligonucleotides of the present disclosure comprise non-naturally occurring nucleotide analogues, e.g., nucleotides which have modified sugar moieties, such as bicyclic nucleotides or 2' modified nucleotides, such as 2' substituted nucleotides. Oligonucleotides are frequently modified at the ribose sugar, primarily with the aim of improving properties such as affinity and/or nuclease resistance. Such modifications include those where the ribose ring structure is modified (e.g. locked nucleic acids or LNAs), where the sugar moiety is replaced by a non-sugar moiety (e.g. peptide nucleic acids or PNAs) or where the substituent groups on the ribose ring are altered to groups other than the hydrogen or 2' and OH group naturally found in DNA and RNA nucleosides.

Non-limiting examples of ring structure modifications are HNAs (hexitol nucleic acids) where the ribose ring is replaced with a hexose ring, an UNA (unlocked nucleic acid) where an unlinked ribose ring lacks a bond between the C2 and C3 carbons or a Locked Nucleic Acid (LNA) where the C2' and C4' of the ribose sugar ring are linked by a methylene bridge (also referred to as a"2'-4' bridge"), which restricts or locks the conformation of the ribose ring. The locking of the conformation of the ribose (also referred to as Bridged Nucleic Acids or BNAs) is associated with an enhanced affinity of hybridization (duplex stabilization) when the LNA is incorporated into an oligonucleotide for a complementary RNA or DNA molecule. Non-limiting examples of LNA nucleosides are beta-D-oxy-LNA, 6'-methyl-beta-D-oxy LNA such as (S)-6'-methyl-beta-D-oxy-LNA (ScET) and 2'-O,4'-C-ethylene-bridged nucleic acid (ENA) or those disclosed in WO 1999/014226, WO 2000/66604, WO 1998/039352, WO 2004/046160, WO 2000/047599, WO 2007/134181 , WO 2010/077578, WO 2010/036698, WO 2007/090071 , WO 2009/006478, WO 2011/156202, WO 2008/154401 , WO 2009/067647, and WO 2008/150729, all of which are herein incorporated by reference in their entireties.

Since BN A modifications enhance both nuclease stability and the affinity of the oligonucleotide for target RNA, they have been incorporated into the flanking regions of gapmers to improve target binding. As such, cEt-flanking 3-10-3 gapmers are more efficacious than the MOE 5-10-5 equivalents. Importantly, BNAs are excluded from the DNA gap region because they are not compatible with RNase H-mediated cleavage. LNA modifications have also been utilized in steric block ASOs, such as miRNA inhibitors.

Non-limiting examples of 2' substituted modified nucleosides are 2'-O-alkyl-RNA, 2'-O-methyl-RNA (2'- OMe), 2'-alkoxy-RNA, 2'-O-methoxyethyl-RNA (MOE), 2'-amino-DNA, 2'-Fluoro-RNA (2'-F), and 2'-F-ANA nucleoside. These modifications increase oligonucleotide nuclease resistance by replacing the nucleophilic 2'-hydroxyl group of unmodified RNA, leading to improved stability in plasma, increased tissue half-lives and consequently prolonged drug effects. These modifications also enhance the binding affinity of the oligonucleotide for complementary RNA and some 2' modifications reduce pro- inflammatory effects. The 2'-ribose modifications are not compatible with RNase H activity, meaning they are typically used for steric block oligonucleotides, or for the flanking sequences in gapmer ASOs. Although most effort was done in modifying the 2' position, substituents can be introduced at the 3', 4' or 5' positions as well.

The present disclosure provides oligonucleotides comprising or consisting of a simple sequence of natural occurring nucleotides - preferably 2'-deoxynucleotides (referred here generally as "DNA"), but also possibly ribonucleotides (referred here generally as "RNA"), or a combination of such naturally occurring nucleotides and one or more non-naturally occurring nucleotides, i.e., "nucleotide analogues", such as nucleotides having the ribose sugar modifications disclosed above.

In some aspects, the oligonucleotide of the present disclosure (e.g., an ASO, a siRNA, or a shRNA) comprises at least two nucleotide analogues. In some aspects, the oligonucleotide of the present disclosure comprises from 3, 4, 5, 6, 7, or 8 nucleotide analogues, e.g. 6 or 7 nucleotide analogues. In some aspects, all the nucleotide analogues are the same. In some aspects, some nucleotide analogs are different. In some aspects, all the nucleotides in the oligonucleotide of the present disclosure are nucleotide analogues, i.e., the oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) is fully modified. In some aspects, when all the nucleotides in the oligonucleotide of the present disclosure are nucleotide analogues, all the nucleotide analogues are the same. In some aspects, when all the nucleotides in the oligonucleotide of the present disclosure are nucleotide analogues, some of the nucleotide analogues are different. In some aspects, all nucleotides in an oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) are 2' modified. In some aspects, all nucleotides in an oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) are 2'-Fluoride and 2'-O-Methyl nucleotides. In some aspects, all nucleotides in an oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) are 2'-Fluoride and 2'-O-methyl nucleotides in an alternating pattern. In some aspects, all nucleotides in a duplex of the present disclosure (e.g., siRNA, or shRNA) are 2'-Fluoride and 2'-O- Methyl nucleotides in an alternating pattern, wherein all or substantially all of the 2'-Fluoride modified nucleotides in the sense strand are complementary to all or substantially all of the 2'-O-methyl modified nucleotides in the antisense strand. In some aspects, a duplex oligonucleotide of the present disclosure (e.g., siRNA, or shRNA) comprises a nucleotide overhang. In some aspects, the nucleotide overhang is a dinucleotide overhang. In some aspects, the dinucleotide overhang is in the antisense strand. In some aspects, the dinucleotide overhang is at the 3' end of the antisense strand. In some aspects, the overhang sequence follows the modification pattern (e.g., alternating pattern) of the rest of the strand. In some aspects, the overhang is complementary to the synaptogyrin-3 mRNA target sequence. In some aspects, the oligonucleotide of the present disclosure has a structure as shown in FIGURE 1.

In some aspects a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises a modification motif set forth in FIGURE 2A or 2B.

In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises a modification motif (e.g., the pattern of distribution of nucleotide analogs along the sense and antisense sequences, internucleoside linkages, conjugate moieties, etc.) disclosed in U.S. Pat. Nos. 8,110,674; 8,420,799; 8,809,516; 9,222,091; 9,708,615; 10,273,477; 9,290,760; 10,233,448; or 9,796,974; U.S. Appl. Publ. No. 2018 and 0258427A1; or Int'l Publ. WO2018098328A1, all of which are herein incorporated by reference in their entireties.

In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one chiral internucleoside linkage. In some aspects, all internucleoside linkages are chiral internucleoside linkages. In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one 8-oxo-deoxyadenosine. In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one phosphoryl DMI amidate diester internucleoside linkage (PN). In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one 8-oxo-deoxyadenosine. In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one phosphoramidite internucleoside linkage. In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one phosphoramidate internucleoside linkage. In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one pseudouridine. In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one isouridine. See, e.g., WO2022/099159 and WO2021/071858, which are herein incorporated by reference in their entireties. In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises at least one glycol nucleic acid (GNA). In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises a loop. In some aspects, a nucleic acid or oligonucleotide of the present disclosure (e.g., an ASO, siRNA, or shRNA) comprises a cleavable loop.

Conjugation

In some aspects, the oligonucleotide of the present disclosure is a conjugate, e.g., a GalNAc conjugate. The delivery potential of ASOs and RNAi molecules such as siRNAs can be enhanced through direct covalent conjugation of various moieties that promote intracellular uptake, target the drug to specific cells/tissues or reduce clearance from the circulation. Non-limiting examples are lipids, peptides, aptamers, antibodies and sugars. Bioconjugates constitute distinct, homogeneous, single-component molecular entities with precise stoichiometry, meaning that high-scale synthesis is relatively simple and their pharmacokinetic properties are well defined. Furthermore, bioconjugates are typically of small size meaning that they generally exhibit favourable biodistribution profiles. For example, conjugating ASOs or siRNAs to the sugar moiety GalNAc results in more productive delivery to hepatocytes without a meaningful shift in distribution to other tissues and results in 15-30 fold increases in potency for RNA targets in those cells.

ASOs and RNAi molecules such as siRNAs can also be loaded to exosomes. Exosomes are heterogeneous, lipid bilayer-encapsulated vesicles approximately 100 nm in diameter that are generated as a result of the inward budding of the multivesicular bodies. Exosomes are thought to be released into the extracellular space by all cells, where they facilitate intercellular communication via the transfer of their complex macromolecular cargoes. Exosomes present numerous favourable properties in terms of oligonucleotide drug delivery of which crossing biological membranes, such as the blood-brain-barrier (BBB) is highly relevant for treatments of CNS disorders. Methods of the application

In one embodiment, the oligonucleotide(s) of the invention such as the RNAi molecule(s) of the invention is man-made and/or is chemically synthesized and/or is typically purified or isolated. Accordingly, the present disclosure provides a method of manufacturing the nucleic acid molecule(s) or the oligonucleotide(s) of the invention comprising chemically synthesizing the nucleic acid molecule(s) or the oligonucleotide(s) of the invention. In some aspects, the method comprises the conjugation of a delivery moiety, e.g., a GalNAc moiety.

The present disclosure also provides a method for designing or manufacturing an oligonucleotide of the present disclosure (e.g., an ASO or a siRNA) capable of inhibiting a human synaptogyrin-3 (hSYNGR3) gene transcript and/or hSYNGR3 protein expression and/or activity in a cell, a tissue, or a subject, wherein the oligonucleotide of the present disclosure is complementary (partially or fully complementary) to any of the target regions of the application as described above. In some aspects, the complementary sequence of the oligonucleotide of the present disclosure comprises or consists of a subsequence of a nucleotide sequence set forth in SEQ ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 69, 82 or 93, mor particularly set forth in SEQ. ID No. 2-4, 6-15, 17-18, 20-21, 23-25, 27-29, 31-41, 43-45, 47- 49, 51-57, 60-68, 70-81 or 83-92. In some aspects, the complementary sequence of the oligonucleotide of the present disclosure partially overlaps of a nucleotide sequence set for in SEQ ID No. 94-249, more particularly in SEQ ID No. 172-249. In some aspects, the complementarity is at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or about 100% complementary.

As used herein the term "manufacturing" refers to chemically synthesizing, e.g., using solid phase synthesis, an oligonucleotide of the present disclosure. In some aspects, manufacturing further comprises chemically attaching or conjugating a moiety such a delivery moiety (e.g., a GalNAc moiety), and/or a targeting moiety.

The present disclosure also provides a method of manufacturing an oligonucleotide of the present disclosure, the method comprising chemically synthesizing the oligonucleotide of the present disclosure using sequential solid phase oligonucleotide synthesis. The present disclosure provides a method of manufacturing an oligonucleotide of the present disclosure comprising a conjugate moiety, wherein the method comprises covalently attaching the conjugate moiety (e.g., at least one non-nucleotide or nonpolynucleotide moiety) covalently to the oligonucleotide disclosed herein. In some aspects, the conjugate moiety (e.g., a non-nucleotide or non-polynucleotide moiety, for example a carbohydrate conjugate moiety such as a GalNAc moiety) is attached to an oligonucleotide disclosed herein directly or via a linker positioned between the oligonucleotide sequence and the conjugate moiety.

In some aspects, the non-nucleotide or non-polynucleotide moiety is a liver targeting moiety that is attached to the 5' -end or to the 3' -end of an oligonucleotide disclosed herein. In some aspects, the liver targeting moiety is linked to the oligonucleotide via a linker. In some aspects, the liver targeting moiety comprises a carbohydrate conjugate moiety comprising a carbohydrate selected from the group consisting of galactose, lactose, N-acetylgalactosamine (GalNAc), mannose, mannose-6-phosphate, and combinations thereof. In some aspects, the carbohydrate conjugate moiety is not a linear carbohydrate polymer. In some aspects, the carbohydrate conjugate moiety is a carbohydrate group comprising 1, 2, 3, or 4 carbohydrate moieties. In some aspects, all the carbohydrate moieties are identical. In some aspects, at least one carbohydrate moiety is different (non-identical) with respect to the other carbohydrate moieties. In some aspects, the carbohydrate conjugate moiety comprises at least one asialoglycoprotein receptor targeting conjugate moiety. In some aspects, the asialoglycoprotein receptor targeting conjugate moiety comprises a monovalent, divalent, trivalent, or tetravalent GalNAc cluster. In some aspects, each GalNAc in the GalNAc cluster is attached to a branch point group via a spacer. In some aspects, the branch point group comprises di-lysine. In some aspects, the spacer comprises a PEG spacer. In some aspects, the linker comprises a C6 to C12 amino alkyl group or a biocleavable phosphate nucleotide linker comprising between 1 to 6 nucleotides.

In some aspects, covalently attaching the conjugate moiety (e.g., a non-nucleotide or non-polynucleotide moiety, such as a GalNAc moiety) to the oligonucleotide comprises: (i) chemically synthesizing the oligonucleotide; and, (ii) adding by chemical synthesis or conjugation the conjugate moiety to the oligonucleotide to yield an oligonucleotide conjugate. In some aspects, adding by chemical synthesis or conjugation the conjugate moiety (e.g., a non-nucleotide or non-polynucleotide moiety, such as a GalNAc moiety) to the oligonucleotide to yield an oligonucleotide conjugate comprises: (i) incorporating by chemical synthesis or conjugation at least one conjugate moiety (e.g., a non-nucleotide or non- polynucleotide moiety, such as a GalNAc moiety) to the oligonucleotide; (ii) incorporating by chemical synthesis or conjugation at least one linker to the oligonucleotide or conjugate moiety (e.g., a non- nucleotide or non-polynucleotide moiety, such as a GalNAc moiety); (iii) incorporating by chemical synthesis or conjugation at least one branching point to the oligonucleotide or conjugate moiety (e.g., a non-nucleotide or non-polynucleotide moiety, such as a GalNAc moiety); (iv) incorporating by chemical synthesis or conjugation at least one spacer to the oligonucleotide or conjugate moiety (e.g., a non- nucleotide or non-polynucleotide moiety, such as a GalNAc moiety); or, (v) a combination thereof. In some aspects, (i) at least one linker is interposed between the oligonucleotide and a branching point; (ii) at least one branching point is interposed between a linker and a conjugate moiety (e.g., a nonnucleotide or non-polynucleotide moiety, such as a GalNAc moiety); (iii) at least one, two, or three conjugate moieties (e.g., a non-nucleotide or non-polynucleotide moiety, such as a GalNAc moiety) are attached to a branching point; (iv) at least one polymer spacer (e.g., a PEG spacer) is interposed between a conjugate moiety (e.g., a non-nucleotide or non-polynucleotide moiety, such as a GalNAc moiety) and a branching point; or, (v) any combination thereof.

Pharmaceutical salt

The oligonucleotides according to the present invention may exist in the form of their pharmaceutically acceptable salts. The term "pharmaceutically acceptable salt" refers to conventional acid-addition salts or base-addition salts that retain the biological effectiveness and properties of the nucleic acid molecules or oligonucleotides of the present invention and are formed from suitable non-toxic organic or inorganic acids or organic or inorganic bases. Acid-addition salts include for example those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, sulfamic acid, phosphoric acid and nitric acid, and those derived from organic acids such as p-toluene sulfonic acid, salicylic acid, methane sulfonic acid, oxalic acid, succinic acid, citric acid, malic acid, lactic acid, fumaric acid, and the like. Base-addition salts include those derived from ammonium, potassium, sodium and, quaternary ammonium hydroxides, such as for example, tetramethyl ammonium hydroxide. The chemical modification of a pharmaceutical compound into a salt is a technique well known to pharmaceutical chemists in order to obtain improved physical and chemical stability, hygroscopicity, flowability and solubility of compounds. It is for example described by Bastin (2000 Organic Process Research & Development 4:427-435) or in Ansel (1995 In: Pharmaceutical Dosage Forms and Drug Delivery Systems, 6th ed., pp. 196 and 1456-1457). For example, the pharmaceutically acceptable salt of the nucleic acid molecules or oligonucleotides provided herein may be a sodium salt. Provided herein is a pharmaceutically acceptable salt of the nucleic acid molecules or oligonucleotides described herein. In one embodiment, the pharmaceutically acceptable salt is a sodium or a potassium salt.

Pharmaceutical Composition

In another aspect, the invention provides pharmaceutical compositions comprising any of the nucleic acid molecules or oligonucleotides described herein or salts thereof and a pharmaceutically acceptable diluent, carrier, salt and/or adjuvant. A pharmaceutically acceptable diluent includes phosphate- buffered saline (PBS) and pharmaceutically acceptable salts include, but are not limited to sodium and potassium salts. In some embodiments the pharmaceutically acceptable diluent is sterile phosphate buffered saline. In some embodiments the nucleic acid molecules or oligonucleotides of the application are used in the pharmaceutically acceptable diluent at a concentration between about 2 and 100 nM, between about 5 and 500 nM, between about 20 and 750 nM, between about 0.05 and 10 pM, between about 1 and 500 pM, between about 2 and 750 pM, between about 0.01 and 1 mM, between about 0.5 and 10 mM or between about 50 and 300 mM solution.

Suitable formulations for use in the present invention are found in Remington's Pharmaceutical Sciences, Mack Publishing Company, Philadelphia, Pa., 17th ed., 1985. For a brief review of methods for drug delivery, see e.g. Langer (1990 Science 249:1527-1533). Non-limiting examples of pharmaceutically acceptable diluents, carriers, adjuvants, suitable dosages, formulations, administration routes, compositions, dosage forms, combinations with other therapeutic agents, pro-drug formulations are provided in W02007/031091, which is herein incorporated by reference in its entirety. The nucleic acid molecules or oligonucleotides of the application or salts thereof may be mixed with pharmaceutically acceptable active or inert substances for the preparation of pharmaceutical compositions or formulations. Compositions and methods for the formulation of pharmaceutical compositions are dependent upon a number of criteria, including but not limited to route of administration, extent of disease, or dose to be administered. Pharmaceutical compositions comprising any of the nucleic acid molecules or oligonucleotides of the application or salts thereof may be sterilized by conventional sterilization techniques or may be sterile filtered. The resulting aqueous solutions may be packaged for use as is or lyophilized, the lyophilized preparation being combined with a sterile aqueous carrier prior to administration.

The pH of the preparations typically will be between 3 and 11, more particularly between 5 and 9 or between 6 and 8, most particularly between 7 and 8, such as 7 to 7.5. The resulting compositions in solid form may be packaged in multiple single dose units, each containing a fixed amount of the nucleic acid molecules or oligonucleotides of the application or salts thereof, such as in a sealed package of tablets or capsules. The composition in solid form can also be packaged in a container for a flexible quantity.

Tauopathic disorders

It was previously shown that SYNGR-3 interacts with pathological Tau at the presynapse and that reducing the level of SYNGR-3 rescued Tau-induced defects in vesicle mobility and neurotransmitter release (Mclnnes et al 2018 Neuron 97:823-835). Accordingly, the present disclosure provides oligonucleotides that are capable of inhibiting the expression and/or activity of synaptogyrin-3 to reduce binding between synaptogyrin-3 and (the N-terminal sequence of) the tau protein. Therefore in a further aspect, any of the nucleic acid molecules or oligonucleotides described in current application is provided for use as a medicament. More particularly for use to treat tauopathies. The present disclosure also provides methods to treatment, prevent, or ameliorate a symptom or sequelae of a disease or disorder disclosed herein comprising administering an effective of any of the nucleic acid molecules or oligonucleotides described herein, or a combination thereof, to a subject in need thereof.

Tauopathies are a diverse group of disorders all having in common their association with prominent accumulation of intracellular tau protein. The tau protein is abundantly expressed in the central nervous system. The group of tauopathies is growing as recently Huntington disease (Fernandez and Nogales et al 2014 Nat Med 20:881-885) and chronic traumatic encephalopathy (CTE; McKee et al 2009 J Neuropathol Exp Neurol 68:709-735) were added.

Different classifications of tauopathies exist. In one classification system, tauopathic disorders are divided in predominant Tau pathologies, tauopathies associated with amyloid deposition and tauopathies associated with another pathology (Williams et al 2006 Intern Med J 36:652-660). Predominant Tau pathologies include progressive supranuclear palsy (PSP), progressive supranuclear palsy-parkinsonism (PSP-P), Richardson's syndrome, argyrophilic grain disease, corticobasal degeneration, Pick's disease, frontotemporal dementia with parkinsonism associated with chromosome 17 (FTDP-17), post-encephalitic parkinsonism, Parkinson's disease complex of Guam, and Guadeloupean parkinsonism. Tauopathic disorders associated with amyloid deposition include Alzheimer's disease, Down's syndrome, dementia pugilistica, familial British dementia and familial Danish dementia. Tauopathic disorders associated with another pathology include myotonic dystrophy, Hallevorden-Spatz disease, and Niemann Pick type C.

Another classification is based on the isoform type found in the aggregates although overlaps may exist: 4R tauopathies include progressive supranuclear palsy (PSP), corticobasal degeneration, tangle predominant dementia, and argyrophilic grain disease. 3R tauopathies include Pick disease, and 3R+4R tauopathies include Alzheimer's disease (Dickson et al 2011 J Mol Neurosci 45:384-389; Murray et al 2014 Alzheimer's Res Ther 6:1). The tau protein is discussed herein in more detail further below.

Further tauopathies include tangle-only dementia, white matter tauopathy with globular glial inclusions, subacute sclerosing panencephalitis, SLC9A6-related mental retardation, non-Guamanian motor neuron disease with neurofibrillary tangles, neurodegeneration with brain iron accumulation, Gerstmann- Straussler-Scheinker disease, frontotemporal lobar degeneration, diffuse neurofibrillary tangles with calcification, chronic traumatic encephalopathy, amyotrophic lateral sclerosis of Guam, amyotrophic lateral sclerosis and parkinsonism-dementia complex, prion protein cerebral amyloid angiopathy, and progressive subcortical gliosis (Murray et al 2014 Alzheimer's Res Ther 6:1; Spillantini & Goedert 2013 Lancet Neurol 12:609-622). Symptoms of tauopathic disorders include clinical or pathological symptoms such as mild cognitive impairment, dementia, cognitive decline (e.g. apathy, impairment in abstract thought), decline of motor function (causing e.g. postural instability, tremor or dystonia), oculomotor and bulbar dysfunction. Criteria for diagnosing dementia are outlined in e.g. the Diagnostic and Statistical Manual of Mental Disorders (DSM) or in the International Classification of Disease (ICD) and are subject to regular updates. The type of clinical symptoms depends on which region of the brain is affected by the tauopathy and explains why Alzheimer's disease is mainly a dementing disease and why Parkinson's disease is mainly affecting movement. Stereotypical temporospatial propagation of tau inclusions creates a consistent pattern of brain lesions in at least Alzheimer's disease and argyrophilic grain disease. The spreading may in part occur in a trans-synaptic manner (Spillantini & Goedert 2013 Lancet Neurol 12:609-622; Liu et al 2012 PloS One 7:e31802). Molecular symptoms of tauopathic disorders include synaptic dysfunction (in particular pre-synaptic dysfunction), neurotoxicity, neuronal degeneration, neuronal dysfunction, synapse loss and amyloid deposition.

Given that the nucleic acid molecules or oligonucleotides herein described are able to target an mRNA encoding synaptogyrin-3 and reducing its expression, any of said nucleic acid molecules or oligonucleotides is thus applicable for use as a medicament. In one embodiment thereto, any of the nucleic acid molecules or oligonucleotides herein described is provided for use in (a method for) treating or inhibiting progression of a tauopathic disorder or for use in (a method for) treating or inhibiting a symptom of a tauopathic disorder. In particular, the nucleic acid molecules or oligonucleotides of the invention are inhibitors of human synaptogyrin-3 expression and/or human synaptogyrin-3 activity. The expression or function of synaptogyrin-3 is (partially) inhibited such as to restore pathological Tau- induced presynaptic dysfunction. In the methods for treating or inhibiting progression of a tauopathic disorder or a symptom of a tauopathic disorder, any of the nucleic acid molecules or oligonucleotides herein described is administered to a subject in need thereof (a subject suffering of or displaying a tauopathy or symptom thereof) in an effective amount, i.e. in an amount sufficient to treat or to inhibit progression of a tauopathic disorder or a symptom of a tauopathic disorder.

For the purpose of treating, preventing or inhibiting (progression of) an intended disease or disorder, and in method for treating, preventing or inhibiting (progression of) an intended disease or disorder, an effective amount of the therapeutic compound is administered to a subject in need thereof. An "effective amount" of an active substance in a composition is the amount of said substance required and sufficient to elicit an adequate response in treating, preventing, inhibiting (progression of) the intended or targeted medical indication. It will be clear to the skilled artisan that such response may require successive (in time) administrations with the composition as part of an administration scheme. The effective amount may vary depending on the nature of the compound, the route of administration of the compound (crossing of the blood-brain barrier and the cell membrane are potential barriers to be taken by oligonucleotides as described herein), the health and physical condition of the individual to be treated, the age of the individual to be treated (e.g. dosing for infants may be lower than for adults) the taxonomic group of the individual to be treated (e.g. human, non-human primate, primate, etc.), the capacity of the individual's system to respond effectively, the degree of the desired response, the formulation of the active substance, the treating doctor's assessment and other relevant factors. The effective amount further may vary depending on whether it is used in monotherapy or in combination therapy. Determination of an effective amount of a compound usually follows from pre-clinical testing in a representative animal or in vitro model (if available) and/or from dose-finding studies in early clinical trials.

Any of the nucleic acid molecules or oligonucleotides described herein is provided for use in (a method for) treating or inhibition progression of a tauopathic disorder wherein the tauopathic disorder is selected from the group consisting of Alzheimer's disease, progressive supranuclear palsy (PSP), progressive supranuclear palsy-parkinsonism (PSP-P), Richardson's syndrome, argyrophilic grain disease, corticobasal degeneration Pick's disease, frontotemporal dementia with parkinsonism associated with chromosome 17 (FTDP-17), post-encephalitic parkinsonism, Parkinson's disease complex of Guam, Guadeloupean parkinsonism, Huntington disease, Down's syndrome, dementia pugilistica, familial British dementia, familial Danish dementia, myotonic dystrophy, Hallevorden-Spatz disease, Niemann Pick type C, chronic traumatic encephalopathy, tangle-only dementia, white matter tauopathy with globular glial inclusions, subacute sclerosing panencephalitis, SLC9A6-related mental retardation, nonGuamanian motor neuron disease with neurofibrillary tangles, neurodegeneration with brain iron accumulation, Gerstmann-Straussler-Scheinker disease, frontotemporal lobar degeneration, diffuse neurofibrillary tangles with calcification, chronic traumatic encephalopathy, amyotrophic lateral sclerosis of Guam, amyotrophic lateral sclerosis and parkinsonism-dementia complex, prion protein cerebral amyloid angiopathy, and progressive subcortical gliosis.

Any of the nucleic acid molecules or oligonucleotides described herein is thus likewise applicable for use in (a method for) treating or inhibition progression of a symptom of tauopathic disorder selected from the group of mild cognitive impairment, dementia, cognitive decline, decline of motor function, oculomotor and bulbar dysfunction, synaptic dysfunction, neurotoxicity, neuronal degeneration, neuronal dysfunction, synapse loss, and amyloid deposition. In particular, in relation to synaptic dysfunction it concerns pre-synaptic dysfunction. Also provided is a method of treating a tauopathic disorder in a subject in need thereof, the method comprising administering comprising administering an effective amount of an oligonucleotide of the present disclosure to the subject. The present disclosure also provides a method of treating or inhibiting progression of a tauopathic disorder or treating or inhibiting a symptom of a tauopathic disorder in a subject in need thereof, the method comprising administering comprising administering an effective amount of an oligonucleotide of the present disclosure to the subject.

"Treatment" refers to any rate of reduction or retardation of the progress of the disease or disorder compared to the progress or expected progress of the disease or disorder when left untreated. More desirable, the treatment results in no/zero progress of the disease or disorder (i.e. "inhibition" or "inhibition of progression") or even in any rate of regression of the already developed disease or disorder. Tauopathies are in general progressive disorders, and progression may imply propagation of pathological tau protein (Asai et al 2015 Nat Neurosci 18:1584-1593; deCalignon et al 2012 Neuron 73:685-697).

"Reduction" or "reducing" of the progress of a disease as used herein refers to a statistically significant reduction. More particularly, a statistically significant reduction upon administering the oligonucleotide of the invention compared to a control situation wherein the oligonucleotide is not administered. In a particular embodiment, said statistically significant reduction is an at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45% or at least about 50% reduction compared to the control situation.

The present disclosure provides an in vitro method of reducing expression levels and/or activity of synaptogyrin-3 in a cell comprising administering an effective amount of an oligonucleotide of the present disclosure to the cell. Also provided is a method of reducing expression levels and/or activity of synaptogyrin-3 in a subject in need thereof comprising administering an effective amount of an oligonucleotide of the present disclosure to the subject. Also provided is method of reducing synaptogyrin-3 levels in a subject in need thereof comprising administering to said subject an effective amount of an oligonucleotide of the present disclosure.

Diagnosis of tauopathic disorders

The nucleic acid molecules or oligonucleotides of the present disclosure can also be used for diagnostic purposes. Magnetic resonance imaging (MRI) in itself allows for radiologic determination of brain atrophy. Midbrain atrophic signs such as the Hummingbird or Penguin silhouette are for instance indicators of progressive supranuclear palsy (PSP). Determination of tau protein content in the cerebrospinal fluid (CSF) may also serve as an indicator of tauopathies. The ratio between the 33 kDa/55 kDa tau-forms in CSF was e.g. found to be reduced in a patients with PSP (Borroni et al 2008 Neurology 71:1796-1803).

Recently, in vivo imaging techniques of neurodegeneration have become available. Such techniques can clearly support the clinical diagnosis of neurodegenerative diseases in general and of tauopathies in particular. In vivo diagnosis of tauopathies benefits from the existence of Tau imaging ligands detectable by positron emission tomography (PET), and include the radiotracers 2-(l-(6-((2-[¹⁸F]fluoroethyl) (methyl) amino)-2-naphthyl)ethylidene) malononitrile ([¹⁸F]FDDNP), 2-(4-aminophenyl)-6-(2-

([¹⁸F]fluoroethoxy))quinolone ([¹⁸F]THK523), and [¹⁸F]T807 and [¹⁸F]T808 (Murray et al 2014 Alzheimer's Res Ther 6:1). In addition, MRI can be used to detect tauopathies, and PET imaging with fluorodeoxyglucose (FDG, ¹⁸F agent) is indicative of synaptic activity (Murray et al 2014 Alzheimer's Res Ther 6:1). Beta-amyloid, that can be detected in vivo, e.g. by using florbetapir (or other amyloid markers) in combination with PET, proved to be an accurate biomarker for at least Alzheimer's disease (Clark et al 2011 J Am Med Assoc 305:275-283) and the florbetapir-PET technique received FDA approval in 2012. The availability of in vivo tauopathy detection techniques is further supportive for selecting subjects that can benefit from synaptogyrin-3 inhibitory therapies as described herein.

Accordingly, the present disclosure provides nucleic acid molecules or oligonucleotides of the present disclosure which are conjugated to a detectable moiety, for example, a radiotracer, a fluorescent moiety (e.g., a fluorescent protein), or any detectable moiety known in the art. Also provided are methods for the diagnosis or prognosis of tauopathic disorders, methods to monitor the efficacy of a treatment, methods to select a patient for treatment, or methods to select a subject for a clinical trial or to exclude a subject from a clinical trial comprising administering a nucleic acid molecule or oligonucleotides of the present disclosure.

Inhibition of synaptogyrin-3

By using the oligonucleotides of the disclosure, inhibition of synaptogyrin-3 is obtained at the expression level. In other words, the administration of an oligonucleotide of the present disclosure can reduce the level of mRNA encoding synaptogyrin-3, which in turn would result in a lower protein expression level of synaptogyrin-3. In some aspects, such reduction of expression levels of synaptogyrin-3 can result in a reduction in synaptogyrin-3 activity. As demonstrated previously (see W02019/016123 and US20220403021A1, which are herein incorporated by reference in their entireties), partial inhibition of synaptogyrin-3 activity is sufficient to restore pathological Tau-induced presynaptic dysfunction. As such, inhibition of synaptogyrin-3 expression and/or activity implies several possible levels of inhibition. In some aspects, the administration of an oligonucleotide of the disclosure can result in a reduction in synaptogyrin-3 mRNA level of at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% with respect to control conditions (e.g., prior to the administration of the oligonucleotide of the present disclosure). In some aspects, the administration of an oligonucleotide of the disclosure can result in a reduction in synaptogyrin-3 protein level of at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% with respect to control conditions (e.g., prior to the administration of the oligonucleotide of the present disclosure). In some aspects, the administration of an oligonucleotide of the disclosure can result in a reduction in synaptogyrin-3 activity level of at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 100% with respect to control conditions (e.g., prior to the administration of the oligonucleotide of the present disclosure).

The skilled person is familiar with multiple ways of determining the level of synaptogyrin-3 in a cell and hence to determine a reduction of the Syngr-3 transcript level compared to a control. A non-limiting example is quantitative reverse transcriptase (RT)-PCR. In current application the Syngr-3 levels have been determined using a TaqMan assay.

Administrating the nucleic acid molecules of the invention

The nucleic acid molecules or the oligonucleotides of the present disclosure can be administered via intravenous, subcutaneous, intramuscular, intracerebral, intracerebroventricular, intraventricular, intraocular, or intrathecal administration. In some embodiments, the administration is via intrathecal administration. "Administering" as used herein, means to give a composition comprising a composition disclosed herein to a subject via a pharmaceutically acceptable route.

SYNGR-3 gene inactivation, i.e. inhibition of expression of the target gene, can be also achieved through the creation of transgenic organisms expressing one of the oligonucleotides of the invention (e.g. siRNA), or by administering said inhibitor to the subject. The nature of the inhibitor (siRNA, shRNA, ASO, etc) and whether the effect is achieved by incorporating the oligonucleotide into the subject's genome or by administering the oligonucleotide is not vital to the invention, as long as said oligonucleotide reduces the level of Syngr-3 transcripts. An oligonucleotide construct can be delivered, for example as an expression plasmid, which when transcribed in the cell, produces the oligonucleotide that is complementary to at least a unique portion of the cellular Syngr-3 RNA. Alternatively, oligonucleotide inhibitors such as siRNA can also be expressed from recombinant circular or linear DNA plasmids using any suitable promoter. Suitable promoters for expressing these inhibitors targeted against Syngr-3 from a plasmid include, for example the U6 or Hl RNA polymerase III promoter sequences and the cytomegalovirus promoter. Selection of other suitable promoters is within the skill in the art. Nonlimiting examples are neuronal-specific promoters, glial cell specific promoters, the human synapsin 1 gene promoter, the Hb9 promotor or the promoters disclosed in US7341847B2.

The recombinant plasmids comprising any of the nucleic acid molecules or oligonucleotides of the invention can also comprise inducible or regulatable promoters for expression of the nucleic acid molecule or oligonucleotide in a particular tissue or in a particular intracellular environment. The nucleic acid molecule or oligonucleotide expressed from recombinant plasmids can either be isolated from cultured cell expression systems by standard techniques, or can be expressed intracellularly, e.g. in brain tissue or in neurons. Nucleic acid molecules or oligonucleotides can also be expressed intracellularly from recombinant viral vectors. The recombinant viral vectors comprise sequences encoding the nucleic acid molecules or oligonucleotides of the invention and any suitable promoter for expressing them. The nucleic acid molecules or oligonucleotides will be administered in an "effective amount" which is an amount sufficient to cause a statistically significant reduction of the Syngr-3 transcript. Generally, an effective amount of a nucleic acid molecule or oligonucleotide targeting Syngr-3 transcripts comprises an intracellular concentration of from about 0.2 nanomolar (nM) to about 100 nM, preferably from about 1 nM to about 10 nM, more preferably from about 2 nM to about 20 nM, more preferably from about 2.5 nM to about 50 nM, even more preferably from about 5 nM to about 75 nM or from about 10 nM to about 150 nM. It is contemplated that greater or lesser amounts of inhibitor can be administered. shRNAs for example can be introduced into the nuclei of target cells using a vector (e.g. bacterial or viral) that optionally can stably integrate into the genome. shRNAs are usually transcribed from vectors, e.g. driven by the Pol III U6 promoter or Hlpromoter. Vectors allow for inducible shRNA expression, e.g. relying on the Tet-on and Tet-off inducible systems commercially available, or on a modified U6 promoter that is induced by the insect hormone ecdysone. A Cre-Lox recombination system has been used to achieve controlled expression in mice. Synthetic shRNAs can be chemically modified to affect their activity and stability. Plasmid DNA or dsRNA can be delivered to a cell by means of transfection (lipid transfection, cationic polymer-based nanoparticles, lipid or cell-penetrating peptide conjugation) or electroporation. Viral vectors include lentiviral, retroviral, adenoviral and adeno-associated viral vectors.

Drug administration across blood-brain barrier

In some aspects, the oligonucleotides of the present disclosure are administered across the blood-brain barrier. The blood-brain barrier (BBB) is a protective layer of tightly joined cells that lines the blood vessels of the brain which prevents entry of harmful substances (e.g. toxins, infectious agents) and restricts entry of (non-lipid) soluble molecules that are not recognized by specific transport carriers into the brain. This poses a challenge in the delivery of drugs, such as the synaptogyrin-3 inhibitors described herein, to the central nervous system/brain in that drugs transported by the blood not necessarily will pass the blood-brain barrier. Although the BBB often is to some degree affected or broken down in case of a tauopathic disorder, it may be needed to rely on a means to enhance permeation of the BBB for a candidate drug for treating a tauopathic disorder to be able to enter the affected brain cells. Thus, in some aspects, the oligonucleotides of the present disclose are formulated, conjugated, or carried by vectors, polymers, cells, or devices, to name a few alternatives, that allow the oligonucleotides to cross the BBB. Several options are nowadays available for delivery of drugs across the BBB (Peschillo et al 2016 J Neurointervent Surg 8:1078-1082; Miller & O' Callaghan 2017 Metabolism 69:S3-S7; Drapeau & Fortin 2015 Current Cancer Drug Targets 15:752-768).

Drugs can be directly injected into the brain (invasive strategy) or can be directed into the brain after BBB disruption with a pharmacological agent (pharmacologic strategy). In some aspects, an oligonucleotide of the present disclosure can be directly injected into the brain, e.g., using a needle or a catheter. In some aspects, an oligonucleotide of the present disclosure can be directed into the brain by BBB disruption with a pharmacological agent. Invasive means of BBB disruption are associated with the risk of hemorrhage, infection or damage to diseased and normal brain tissue from the needle or catheter. Direct drug deposition may be improved by the technique of convection-enhanced delivery. Accordingly, in some aspects, an oligonucleotide of the present disclosure can be administered via convection- enhanced delivery.

Longer term delivery of a therapeutic protein (e.g. a neurotrophic factor or nerve growth factor, or a proteinaceous synaptogyrin-3 inhibitor as described herein) can be achieved by implantation of genetically modified stem cells, by recombinant viral vectors, by means of osmotic pumps, or by means of incorporating the therapeutic drug in a polymer (slow release; can be implanted locally). Thus, in some aspects, an oligonucleotide of the present disclosure can be administered, e.g., by implantation of genetically modified cells (e.g., stem cells), recombinant vectors (e.g., viral vectors), delivery devices (e.g., pumps such as osmotic pumps), or incorporation in a polymer.

Pharmacologic BBB disruption has the drawback of being non-selective and can be associated with unwanted effects on blood pressure and the body's fluid balance. This is circumvented by targeted or selective administration of the pharmacologic BBB disrupting agent. As an example, intra-arterial cerebral infusion of an antibody (bevacizumab) in a brain tumor was demonstrated after osmotic disruption of the BBB with mannitol (Boockvar et al. 2011, J Neurosurg 114:624-632); other agents capable of disrupting the BBB pharmacologically include bradykinin and leukotriene C4 (e.g. via intracarotid infusion; Nakano et al. 1996, Cancer Res 56:4027-4031). Thus, in some aspects, the oligonucleotides of the present disclosure are formulated in combination with a pharmacologic BBB disrupting agent. In some aspects, the oligonucleotides of the present disclosure are administered in combination with a pharmacologic BBB disrupting agent. In some aspects, the pharmacologic BBB disrupting agent is administered prior to the administration of the oligonucleotide of the present disclosure. In some aspects, the pharmacologic BBB disrupting agent is administered concurrently to the administration of the oligonucleotide of the present disclosure. In some aspects, the pharmacologic BBB disrupting agent is administered subsequently to the administration of the oligonucleotide of the present disclosure. In some aspects, the pharmacologic BBB disrupting agent comprises mannitol, bradykinin, leukotriene C4, or a combination thereof.

BBB transcytosis and efflux inhibition are other strategies to increase brain uptake of drugs supplied via the blood. Using transferrin or transferrin-receptor antibodies as carrier of a drug is one example of exploiting a natural BBB transcytosis process (Friden et al. 1996, J Pharmacol Exp Ther 278:1491-1498). Exploiting BBB transcytosis for drug delivery is also known as the molecular Trojan horse strategy. In some aspects, the oligonucleotides of the present disclosure are conjugated to carrier, e.g., transferrin or a transferrin-receptor antibody. In some aspects, the oligonucleotides of the present disclosure are conjugated or formulated to transverse the BBB via transcytosis. Another mechanism underlying BBB, efflux pumps or ATP-binding cassette (ABC) transporters (such as breast cancer resistance protein (BCRP/ABCG2) and P-glycoprotein (Pgp/MDRl/ABCBl)), can be blocked in order to increase uptake of compounds (e.g. Carcaboso et al. 2010, Cancer Res 70:4499-4508). In some aspects, the oligonucleotides of the present disclosure can be formulated in combination with a compound that can block an ABC transporter, a compound that can block P-glycoprotein, or a combination thereof.

Kumar et al (2007 Nature 448:39-43) demonstrated uptake of siRNAs in the brain after coupling to a 29- amino acid peptide derived from rabies virus glycoprotein (RVG) which is specifically binding the acetylcholine receptor. In some aspects, the oligonucleotides of the present disclosure are conjugated to RVG.

Therapeutic drugs can alternatively be loaded in liposomes to enhance their crossing of the BBB, an approach also known as liposomal Trojan horse strategy. Thus, in some aspects, the oligonucleotides of the present disclosure are formulated in liposomes, e.g., liposomes for use in a liposomal Trojan horse strategy.

Especially in the field of treating cognitive and neurodegenerative disorders there has been quite some interest in intranasal delivery of drugs (e.g. Muhs et al. 2007, Proc Natl Acad Sci USA 104:9810-9815; Kao et al. 2000, Pharm Res 17:978-984; Hanson & Frey 2008, BMC Neurosci 9 (Suppl3): 55). This strategy is based on the trigeminal and olfactory nerves that innervate the nasal epithelium, representing direct connections between the external environment and the brain. Thus, in some aspects, the oligonucleotides of the present disclosure are formulated for intranasal delivery.

A more recent and promising avenue for delivering therapeutic drugs to the brain consists of (transient) BBB disruption by means of ultrasound, more particularly focused ultrasound (FUS; Miller et al. 2017, Metabolism 69:S3-S7). Besides being non-invasive, this technique has, often in combination with realtime imaging, the advantage of precise targeting to a diseased area of the brain. Therapeutic drugs can be delivered in e.g. microbubbles e.g. stabilized by an albumin or other protein, a lipid, or a polymer. Therapeutic drugs can alternatively, or in conjunction with microbubbles, be delivered by any other method, and subsequently FUS can enhance local uptake of any compound present in the blood (e.g. Nance et al. 2014, J Control Release 189:123-132). Just one example is that of FUS-assisted delivery of antibodies directed against toxic amyloid-beta peptide with demonstration of reduced pathology in mice (Jordao et al. 2010, PloS One 5:el0549). Microbubbles with a therapeutic drug load can also be induced to burst (hyperthermic effect) in the vicinity of the target cells by means of FUS, and when driven by e.g. a heat shock protein gene promoter, localized temporary expression of a therapeutic protein can be induced by ultrasound hyperthermia (e.g. Lee Titsworth et al. 2014, Anticancer Res 34:565-574). Alternatives for ultrasound to induce the hyperthermia effect are microwaves, laser-induced interstitial thermotherapy, and magnetic nanoparticles (e.g. Lee Titsworth et al. 2014, Anticancer Res 34:565-574). Thus, in some aspects, the oligonucleotides of the present disclosure are formulated for FUS-mediated delivery. Intracellular drug administration

In some aspects, the oligonucleotides of the present disclosure are formulated for intracellular administration. Besides the need to cross the BBB, drugs targeting disorders of the central nervous system, such as the synaptogyrin-3 inhibitors described herein, may also need to cross the cellular barrier. Although most antisense oligonucleotides are readily taken up by neurons and glia after reaching the nervous system, it can be advantageous to use facilitators of intracellular drug uptake.

One solution is the use of cell-penetrating proteins or peptides (CPPs). Such peptides enable translocation of the drug of interest coupled to them across the plasma membrane. CPPs are alternatively termed Protein Transduction Domains (TPDs), usually comprise 30 or less (e.g. 5 to 30, or 5 to 20) amino acids, and usually are rich in basic residues, and are derived from naturally occurring CPPs (usually longer than 20 amino acids), or are the result of modelling or design. A non-limiting selection of CPPs includes the TAT peptide (derived from HIV-1 Tat protein), penetratin (derived from Drosophila Antennapedia -Antp), pVEC (derived from murine vascular endothelial cadherin), signal-sequence based peptides or membrane translocating sequences, model amphipathic peptide (MAP), transportan, MPG, polyarginines; more information on these peptides can be found in Torchilin 2008 (Adv Drug Deliv Rev 60:548-558) and references cited therein. The commonly used CPP is the transduction domain of TAT termed TATp. The TAT peptide was e.g. used to shuffle a tau-fragment into neuronal cells (Zhou et al. 2017).

CPPs can be coupled to carriers such as nanoparticles, liposomes, micelles, or generally any hydrophobic particle. Coupling can be by absorption or chemical bonding, such as via a spacer between the CPP and the carrier. To increase target specificity an antibody binding to a target-specific antigen can further be coupled to the carrier (Torchilin 2008, Adv Drug Deliv Rev 60:548-558)

CPPs have already been used to deliver payloads as diverse as plasmid DNA, oligonucleotides, siRNA, peptide nucleic acids (PNA), proteins and peptides, small molecules and nanoparticles inside the cell (Stalmans et al. 2013, PloS One 8:e71752).

Kits and products of manufacture

Also provided herein are kits and products of manufacture comprising one or more compositions (e.g., an oligonucleotide of the present disclosure or pharmaceutical compositions comprising an oligonucleotide of the present disclosure) described herein. In some aspects, provided herein is a pharmaceutical pack or kit comprising one or more containers filled with one or more of the ingredients of the pharmaceutical compositions described herein. In some aspects, the kit or product of manufacture comprises, e.g., a first container comprising a first pharmaceutical composition comprising an oligonucleotide of the present disclosure, a second container containing a solvent, and optionally an instruction for use.

In some aspects, the kit or product of manufacture comprises a container comprising an oligonucleotide of the present disclosure and optionally an instruction for use.

In some aspects, the kit contains a pharmaceutical composition described herein and any prophylactic or therapeutic agent, such as those described herein. In some aspects, the kit further comprises instructions to administer a composition of the present disclosure according to any method disclosed herein. In some aspects, the kit is for use in the treatment of a medical indication disclosed herein. In some aspects, the kit is a diagnostic kit.

All of the references cited above, as well as all references cited herein, are incorporated herein by reference in their entireties.

The sequences of biomolecules (e.g., proteins, genes) disclosed herein and identified by either database accession number or gene name are incorporated by reference. The database accession numbers disclosed herein (e.g. GenBank accession numbers) refer to the database version that in effect on February 1, 2023. The nucleic acid sequences of genes identified by name as well as their official names and alternative names correspond to those in the version of the GenBank database active on February 1, 2023, and are herein incorporated by reference. The amino acid sequences of proteins identified by name or translation products of genes identified by name as well as their official and alternative names correspond to those in the version of the UniProt database active on February 1, 2023, and are herein incorporated by reference.

Although the invention has been described and illustrated in the foregoing illustrative embodiments, it is understood that the present disclosure has been made only by way of example, and the numerous changes in the details of implementation of the invention can be made without departing from the spirit and scope of the invention, which is only limited by the claims that follow. Features of the disclosed embodiments can be combined and rearranged in various ways. The various embodiments can be combined with one or more other embodiments to form new embodiments.

The following examples are offered by way of illustration and not by way of limitation. EXAMPLES

Example 1. Design and synthesis of oligonucleotides

A bioinformatic analysis was performed to identify potential screening candidate siRNAs targeting human SYNGR3 mRNA (NCBI gene ID: 9143). The bioinformatical approach assumed a canonical siRNA structure. Positions 2 and 18 (5' -3') of the sense and antisense strand were used for the specificity calculations. Positions 1-19 (5' -3') of the antisense strand were used to assess cross reactivity and human SNP analysis.

The following parameters were assessed:

(a) Species cross reactivity for human, cynomolgus monkey, rhesus monkey and mouse siRNAs were selected that target at least all protein-coding transcripts of the target gene and for each species. The analysis was based on a canonical siRNA design using 19 bases and 17 bases (without considering positions 1 and 19) for cross reactivity. We considered a 17-mer (position 2-18) perfect match and possibly single mismatch hits with target sequences in the secondary species.

(b) Specificity

Target specificity in human, rhesus monkey, cynomolgus monkey and mouse was performed to identify siRNAs with lowest sequence complementary to any non-target transcript. Our analysis considered the likelihood of unintended downregulation of any other transcript by full or partial complementarity of a siRNA strand (up to 4 mismatches in within position 2-18; it was based on the number and position of the mismatches and described the predicted most likely off-target(s) for antisense and sense strand of each siRNA.

(c) microRNA-like off-target effects

Identicalness of siRNA seed region and seed region of known miRNAs (microRNAs) is an important feature since siRNAs can function in a miRNA-like manner via base-pairing with complementary sequences within the 3'-UTR of mRNA molecules. To circumvent that siRNAs would act via functional miRNA binding sites, we avoided siRNA strands, that contained natural miRNA seed regions (position 2-7).

(d) Single-nucleotide polymorphisms binding

Analysis of human SNP database (NCBI DB SNP) was performed to identify siRNAs targeting regions with known SNPs. When data was available, we also included positions of SNPs within the target sequence as well as minor allele frequency. All target sites with abundant SNPs were excluded.

(e) Seguence activity based on current algorithms siRNA activity prediction was done based on canonical siRNA design.

(f) Identify molecules that can silence specific SyngrS transcripts Considering all the parameters, we selected 192 siRNAs sequences to be synthetized and tested in cell lines.

The siRNAs molecules were fully modified with 2'-Fluoride and 2'-O-methyl as shown in FIGURE 1.

Example 2. Dual-dose screening

We seeded SH-SY5Y cells (ATCC, CRL-2266) at a density of 20.000 cells/well on collagen-coated 96-well tissue culture plates, followed by transfection of cells using Dharmafect-4 (0.5 pl/well; Horizon Discovery). siRNAs were added to cells at final concentrations of 20 nM and 2 nM and assay plates were incubated for 24h at 37°C / 5% CO2 in a humidified incubator. To monitor on-target mRNA expression levels we have used the branched DNA (bDNA) technology. The bDNA assay provides a unique and powerful tool for reliable quantification of nucleic acid molecules. The bDNA assay directly measures nucleic acid molecules at physiological levels by boosting the reporter signal, rather than replicating target sequences as the means of detection, and hence avoids the errors inherent in the extraction and amplification of target sequences.

Upon release of the target RNA, several oligonucleotide probes were incubated to allow binding to Syngr3 (and GAPDH as a control). Probes for Syngr3 were custom and made by ThermoFisher Scientific, Assay ID: DRAAACA). Addition of a fluorescent reporter generates a signal directly proportional to the amount of target RNA present in the sample.

Some of the tested siRNAs showed toxicity in the assay, visible as a drop in mean value of hsGAPDH mRNA expression levels. The mean ratio of on-target hsSYNGR3/hsGAPDH was artificially set to 100% and used to normalize all other samples. All data were generated in quadruplicates. Results of the dualdose screen are shown in TABLE 1. As can be observed from TABLE 1, when we selected for a reduced synaptogyrin-3 expression level of at least 15% in the 2 nM or 20 nM concentration, regions within the synaptogyrin-3 emerged as target regions for designing antisense and/or RNAi molecules capable of reducing the expression of syngr-3 in a cell, tissue or subject. A first target region is built up by the siRNA molecules R2000060, R2000061 and R2000064. A second target region by R2000072, R2000073, R2000075, R2000076, R2000077, R2000080, R2000081, R2000083, R2000084 and R2000085. A third target region by R2000095 and R2000097, a fourth target region by R2000099 and R2000100 and a fifth target region by R2000104, R2000105 and R2000106. A sixth target region comprises oligonucleotides R2000123, R2000124 and R2000125. A seventh target region is formed by R2000130, R2000132, R2000133, R2000134, R2000135, R2000136, R2000137, R2000138, R2000140, R2000141 and R2000142. An eight target region by R2000156, R2000157 and R2000158 and finally two larger target regions defined by R2000175, R2000176, R2000177, R2000179, R2000180, R2000181, R2000182, R2000183, R2000187 and R2000190, and defined by R2000191-R2000221.

From the siRNA molecules that make up the target regions, we further selected 48 siRNAs molecules to be tested in a dose-response curve. The selection was based on activity, target sequence binding, and cross-reactivity.

Example 3. Dose-response curve

We seeded SH-SY5Y cells at a density of 20.000 cells/well on collagen-coated 96-well tissue culture plates, followed by transfection of cells using Dharmafect-4 (0.5 pl/well). For the transfection the highest siRNA concentration tested was 100 nM. Using 2-fold dilution steps and 10 data points, the lowest final siRNA concentration tested was 0.2 nM. Cells were incubated for 48h at 37°C/5% CO2 in a humidified incubator, followed by cell lysis and bDNA analysis to monitor on- target mRNA expression levels relative to hsGAPDH mRNA levels. All data were generated in quadruplicates. Results of the dose-response curve (DRC) are shown in TABLE 1-2.

EXPERIMENTAL PROCEDURES

Measuring Syngr3 levels

SH-SY5Y cells were incubated for 24 hours with the siRNAs to be tested. Media was removed and the SH- SY5Y cells (sourced from ATCC, CRL-2266) were lysed by addition of 150pl lysis mixture (1 volume lysis mixture, 2 volumes nuclease-free water) per 96-well and by subsequent incubation at 53°C for at least 60 minutes. Upon release of the target RNA, several oligonucleotide probes were incubated to allow binding to Syngr3 (and GAPDH as a control). Probes for Syngr3 were custom and made by ThermoFisher Scientific, Assay ID: DRAAACA). During this incubation, the probes cooperatively hybridize to the Syngr3. 50pl working probe set hsSYNGR3 (gene target, Synaptogyrin 3 from Homo sapiens) and 90pl working probe set hsGAPDH (endogenous control, Glyceraldehyde-3-phosphate dehydrogenase from Homo sapiens), and 50pl (for hsSYNGR3) and lOpI (for hsGAPDH) of cell lysate were then added to the capture plates provided by the manufacturer. Capture plates were incubated at 53°C for approximately 16-20 hours. The next day, the capture plates were washed 3 times with at least 300pl of lx Wash Buffer (nuclease-free water, wash buffer component 1 and wash buffer component 2). lOOpI of pre-amplifier working reagent was added to both hsSYNGR3 and hsGAPDH capture plates, which were sealed with clear adhesive foil and incubated for 1 hour at 53°C. Following incubation, the wash step was repeated, then lOOpI amplifier working reagent was added to both hsSYNGR3 and hsGAPDH capture plates. After 1 hour incubation at 53°C, the wash and dry steps were repeated, and lOOpI label probe was added per 96-well to all capture plates. Capture plates were incubated for 53°C for 1 hour. The plates were then washed with lx wash buffer and dried, and then 100 .l substrate was added to the capture plates sealed by adhesive aluminum foil. Following 30 minutes of incubation in the dark, luminescence was read using 1420 Luminescence Counter (WALLAC VICTOR Light, Perkin Elmer, Rodgau-Jugesheim, Germany).

For each siRNA, four wells with SH-SY5Y cells were transfected in parallel, and individual data points were collected from each well separately. For each well, the hsSYNGR3 mRNA level was normalized to the hsGAPDH mRNA level. The activity of a given hsSYNGR3 targeting siRNA was expressed as percent hsSYNGR3 mRNA concentration (normalized to hsGAPDH mRNA) in treated cells, relative to the hsSYNGR3 mRNA concentration (normalized to hsGAPDH mRNA) averaged across control wells.

Claims

1. An oligonucleotide of 10 to 70 nucleotides in length, comprising a contiguous nucleotide sequence of at least 10 contiguous nucleotides in length, the contiguous nucleotide sequence being at least 90% complementary to an equal length portion of a target region within the Synaptogyrin-3 transcript as depicted in SEQ ID No. 1, wherein the target region is comprised between nucleobase 205 and 265, 255 and 348, 338 and 387, 369 and 433, 422 and 531, 603 and 656, 641 and 714, 717 and 768, 1150 and 1600, 1743 and 1868 or between nucleobase 1865 and 2026 of SEQ. ID No. 1 and wherein the endpoints are included.

2. The oligonucleotide according to claim 1, wherein the oligonucleotide can bind to the Synaptogyrin- 3 transcript as depicted in SEQ ID No. 1.

3. The oligonucleotide according to any of the previous claims, wherein the oligonucleotide is capable of statistically significantly reducing the level of Synaptogrin-3 transcript in a cell compared to a control condition in the absence of the oligonucleotide.

4. The oligonucleotide according to any of the previous claims, wherein the oligonucleotide is a double stranded nucleic acid molecule.

5. The oligonucleotide according to claim 4, wherein the double stranded oligonucleotide is an RNAi molecule or an RNA duplex.

6. The oligonucleotide according to claim 5, wherein the RNAi molecule is an siRNA, a divalent siRNA or a shRNA.

7. The oligonucleotide according to any of claims 1-3, wherein the oligonucleotide is a single stranded nucleic acid molecule.

8. The oligonucleotide according to claim 7, wherein the single stranded oligonucleotide is the antisense portion of an RNAi molecule.

9. The oligonucleotide according to any of claims 4-8, wherein the sense and/or antisense strand comprises between 15 and 25 nucleotides in length.

10. The oligonucleotide according to any of claims 4-9, wherein the antisense strand is 21 nucleotides in length.

11. The oligonucleotide according to any of the previous claims comprising at least one single stranded nucleotide overhang.

12. The oligonucleotide according to any of the previous claims, wherein said target region is selected from SEQ ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 82 or 93.

13. The oligonucleotide according to any of the previous claims, wherein said target region is selected from the group consisting of SEQ ID No. 2-4, 6-15, 17-18, 20-21, 23-25, 27-29, 31-41, 43-45, 47-49, 51-57, 70-81, and 90-92.

14. The oligonucleotide according to any of the previous claims wherein the contiguous nucleotide sequence of at least 10 contiguous nucleotides in length shows at least 90% sequence identity to any of SEQ. ID No. 172-218, 228-239 or 247-249.

15. The oligonucleotide according to any of the previous claims, wherein the oligonucleotide comprises one or more internucleoside linkage and/or one or more 2' sugar modified nucleosides.

16. The oligonucleotide according to claim 15 wherein the internucleoside linkage is a phosphorothioate internucleoside linkage and/or wherein the 2' sugar modified nucleoside is selected from the group consisting of 2'-O-methyl-, 2'-O-methoxyethy-, 2'-O-alkyl-, 2' -alkoxy, 2' -amino-, 2'-fluoro- and LNA nucleosides.

17. The oligonucleotide according to claim 15 wherein all oligonucleosides are modified with a phosphorothioate internucleoside linkage or with a 2'-O-methyl group.

18. A pharmaceutical composition comprising the oligonucleotide according to any of the preceding claims.

19. The oligonucleotide according to any of claims 1-17 or the pharmaceutical composition according to claim 18 for use as a medicament.

20. The oligonucleotide according to any of claims 1-17 or the pharmaceutical composition according to claim 18 for use in treating or inhibiting progression of a tauopathic disorder or for use in treating or inhibiting a symptom of a tauopathic disorder.

21. The oligonucleotide according to any of claims 1-17 or the pharmaceutical composition according to claim 18 for use according to claim 20 wherein the tauopathic disorder is selected from the group consisting of Alzheimer's disease, progressive supranuclear palsy (PSP), progressive supranuclear palsy-parkinsonism (PSP-P), Richardson's syndrome, argyrophilic grain disease, corticobasal degeneration Pick's disease, frontotemporal dementia with parkinsonism associated with chromosome 17 (FTDP-17), post-encephalitic parkinsonism, Parkinson's disease complex of Guam, Guadeloupean parkinsonism, Huntington disease, Down's syndrome, dementia pugilistica, familial British dementia, familial Danish dementia, myotonic dystrophy, Hallevorden-Spatz disease, Niemann Pick type C, chronic traumatic encephalopathy, tangle-only dementia, white matter tauopathy with globular glial inclusions, subacute sclerosing panencephalitis, SLC9A6-related mental retardation, non-Guamanian motor neuron disease with neurofibrillary tangles, neurodegeneration with brain iron accumulation, Gerstmann-Straussler-Scheinker disease, frontotemporal lobar degeneration, diffuse neurofibrillary tangles with calcification, chronic traumatic encephalopathy, amyotrophic lateral sclerosis of Guam, amyotrophic lateral sclerosis and parkinsonism-dementia complex, prion protein cerebral amyloid angiopathy, and progressive subcortical gliosis.

22. The oligonucleotide according to any of claims 1-17 or the pharmaceutical composition according to claim 18 for use according to claim 20 wherein the symptom of the tauopathic disorder is selected from the group of mild cognitive impairment, dementia, cognitive decline, decline of motor function, oculomotor and bulbar dysfunction, synaptic dysfunction, neurotoxicity, neuronal degeneration, neuronal dysfunction, synapse loss, and amyloid deposition.

23. The oligonucleotide according to any of claims 1-17 or the pharmaceutical composition according to claim 18 for use according to claim 22 wherein the synaptic dysfunction is pre-synaptic dysfunction.

24. A nucleic acid sequence selected from SEQ. ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 82 or 93 for designing an antisense or RNAi molecule capable of reducing the level of Synaptogyrin-3 transcript in a cell with at least 15% compared to a control situation in the absence of said antisense or RNAi molecule.

25. A method of treating or inhibiting progression of a tauopathic disorder or treating or inhibiting a symptom of a tauopathic disorder comprising administering an effective dose of a oligonucleotide according to any of claims 1 and 17 or the pharmaceutical composition according to claim 18 to a subject in need thereof.

26. The method of claim 25, wherein the tauopathic disorder is selected from the group consisting of Alzheimer's disease, progressive supranuclear palsy (PSP), progressive supranuclear palsyparkinsonism (PSP-P), Richardson's syndrome, argyrophilic grain disease, corticobasal degeneration Pick's disease, frontotemporal dementia with parkinsonism associated with chromosome 17 (FTDP- 17), post-encephalitic parkinsonism, Parkinson's disease complex of Guam, Guadeloupean parkinsonism, Huntington disease, Down's syndrome, dementia pugilistica, familial British dementia, familial Danish dementia, myotonic dystrophy, Hallevorden-Spatz disease, Niemann Pick type C, chronic traumatic encephalopathy, tangle-only dementia, white matter tauopathy with globular glial inclusions, subacute sclerosing panencephalitis, SLC9A6-related mental retardation, non-Guamanian motor neuron disease with neurofibrillary tangles, neurodegeneration with brain iron accumulation, Gerstmann-Straussler-Scheinker disease, frontotemporal lobar degeneration, diffuse neurofibrillary tangles with calcification, chronic traumatic encephalopathy, amyotrophic lateral sclerosis of Guam, amyotrophic lateral sclerosis and parkinsonism-dementia complex, prion protein cerebral amyloid angiopathy, and progressive subcortical gliosis.

27. The method of claim 25, wherein the symptom of the tauopathic disorder is selected from the group of mild cognitive impairment, dementia, cognitive decline, decline of motor function, oculomotor and bulbar dysfunction, synaptic dysfunction, neurotoxicity, neuronal degeneration, neuronal dysfunction, synapse loss, and amyloid deposition.

28. The method of claim 27, wherein the synaptic dysfunction is pre-synaptic dysfunction.

29. An antisense oligonucleotide or RNAi molecule capable of reducing the level of synaptogyrin-3 mRNA, synaptopgyrin-3 protein, synaptogyrin-3 activity, or a combination thereof in a cell by least 15% compared to a control situation in the absence of said antisense or RNAi molecule, wherein the antisense oligonucleotide or RNAi molecule nucleic acid sequence targets a subsequence of an mRNA encoding synaptogyrin-3 selected from the group consisting of SEQ. ID No. 5, 16, 19, 22, 26, 30, 42, 46, 50, 58, 59, 82 and 93.