Antisense oligonucleotides to meg3

Abstract

The present invention relates to oligonucleotides for reducing the level of MEG3 expression in a cell. The present invention further relates to derived products (vectors, virus particles, conjugates, delivery particles, pharmaceutical compositions), the use of the oligonucleotides and derived products as medicaments, as well as methods of treatment or prevention of diseases and particularly of neurodegenerative diseases, cardiovascular diseases, lung disease and diabetes. The oligonucleotides are effective at reducing the level of MEG3 expression in a cell and thus provide effective treatment options for related diseases.

Description

[0001] ANTISENSE OLIGONUCLEOTIDES TO MEG3

[0002] FIELD OF THE INVENTION

[0003] The present invention relates to oligonucleotides for reducing the level of MEG3 expression in a cell. The present invention further relates to derived products (vectors, virus particles, delivery particles, oligonucleotide conjugates, pharmaceutical compositions), methods of reducing the level of MEG3 expression in a cell, methods of treatment or prevention of neurodegenerative diseases, cardiovascular diseases, lung disease, diabetes, and methods of inhibiting necroptosis.

[0004] BACKGROUND TO THE INVENTION

[0005] Oligonucleotides are short DNA and / or RNA molecules, typically several tens of nucleotides or base pairs long, which can be used to regulate gene expression in a target-specific manner and therefore form the basis of a number of therapeutic modalities. Antisense oligonucleotides (ASOs), short interfering RNAs (siRNAs), and short hairpin RNAs (shRNAs) are some examples of well understood and widely used forms of oligonucleotides for modulating gene expression. Where gene expression is reduced, this is known as gene silencing or knockdown. Such nucleic acid therapeutics typically work by binding to targets of interest such as target genes or RNA transcripts through base pair complementarity.

[0006] ASOs are short, single-stranded oligonucleotides, that are able to bind to RNA transcripts of genes in a sequence-specific manner via Watson-Crick base pairing. Where an ASO comprises DNA, sequence-specific binding of an ASO to a target RNA transcript creates a DNA: RNA hybrid, leading to the recruitment of RNase H which recognises DNA: RNA hybrids. RNase H cleaves the duplex structure, causing target RNA degradation and gene silencing. ASO binding may also modify gene expression by, for example, blocking RNA translation. ASOs are often chemically modified to increase their stability, target binding affinity, bioavailability and cellular uptake. siRNAs are double-stranded RNA molecules that are incorporated into a cell’s endogenous RNA-induced silencing complex (RISC), which unwinds the siRNA into a sense and an antisense strand and uses one of these strands to target, cleave and degrade complementary target RNA transcripts causing gene silencing. shRNAs are RNA molecules that form hairpin structures that are processed within a cell into siRNAlike fragments. These fragments are then incorporated into RISC, leading to gene silencing through target RNA transcript degradation. Oligonucleotide-based technologies represent cutting-edge advancements in the field of targeted, personalised therapies. Maternally Expressed Gene 3 (MEG3) is a long noncoding RNA (IncRNA) which regulates cellular functions including cell viability and apoptosis, and is over-expressed in many diseases including cancer retinopathy, osteoarthritis and cardiovascular disease (Bi et al., 2020). MEG3 is implicated in diseases including cardiovascular diseases such as cardiac fibrosis (Piccoli et al. 2017), lung diseases such as obstructive pulmonary disease (Bi et al., 2020), neurodegenerative diseases such as Alzheimer’s disease (Balusu et al. 2023), and diabetes (Zhang, 2019). MEG3 over-expression is known to induce necroptosis in neurons and thus contributes to the progression of neurodegenerative diseases such as Alzheimer’s disease (Balusu et al. 2023). Given the high relevance of MEG3 to a number of different diseases, nucleic-acid based therapeutic agents are needed that effectively target MEG3.

[0007] Several challenges can hinder the development of oligonucleotides which effectively modify gene expression. First, it is not always clear how many isoforms (i.e. splice variants) of a target RNA transcript exist, and if multiple isoforms do exist, which one or more isoforms and specifically which exons comprised within these one or more isoforms should be targeted to effectively modify gene expression. This is particularly pertinent to targeting long non-coding RNAs, which are typically multi-exonic and highly spliced transcripts. Second, it is difficult to predict which specific sites in a target RNA transcript lead to substantial reductions in gene expression when targeted using oligonucleotides. Biological complexities affecting this include for example sequence-dependent secondary structure formation of the target RNA transcript or the oligonucleotide itself which may affect the availability of specific sequence sites for binding, and the occupancy of certain sites of the target RNA transcript by RNA binding proteins which may prevent those sites from being accessible to oligonucleotides. Third, effective intracellular delivery, tissue targeting, stability against degradation, and bioavailability all impact the extent to which oligonucleotides are able to modify gene expression. Fourth, unwanted effects such as cytotoxicity, which could arise for example by oligonucleotides either eliciting an immune response or binding to undesired off-target locations, are also important challenges to overcome. Balancing oligonucleotide chemical modifications to enhance stability and bioavailability without increasing cytotoxicity is a crucial consideration. Overcoming these challenges to develop oligonucleotides that can effectively modify gene expression requires extensive research and optimisation.

[0008] It would be useful to develop oligonucleotides capable of effectively modifying, for example reducing, the level of MEG3 expression, preferably without any unwanted side-effects such as cytotoxicity. It would be even more useful if, given that the long-non-coding RNA that MEG3 encodes has multiple different isoforms, a range of such oligonucleotides were identified, in order to provide a pool of potential candidates for developing lead therapeutics. It is an object of one or more aspects or embodiments of the present invention to address one or more problems in the art such as those mentioned above.

[0009] SUMMARY OF THE INVENTION

[0010] According to a first aspect of the present invention, there is provided an oligonucleotide for reducing the level of MEG3 expression in a cell, wherein the oligonucleotide is capable of binding to a target sequence at least partially (or wholly) within a region of a MEG3 transcript selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 3.

[0011] In a second aspect, there is provided a vector comprising a nucleic acid encoding the oligonucleotide of the first aspect.

[0012] In a third aspect, there is provided a virus particle comprising the oligonucleotide of the first aspect or the vector of the second aspect.

[0013] In a fourth aspect, there is provided a conjugate comprising the oligonucleotide of the first aspect, or the vector of the second aspect, covalently linked to a delivery group, preferably wherein the delivery group is a cell penetrating peptide.

[0014] In a fifth aspect, there is provided delivery particle comprising the oligonucleotide of the first aspect, vector of the second aspect, or the conjugate of the fourth aspect, preferably wherein the delivery particle is a nanoparticle or a vesicle, more preferably a lipid nanoparticle (LNP).

[0015] In a sixth aspect, there is provided a pharmaceutical composition comprising the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, or delivery particle of the fifth aspect, and one or more pharmaceutically acceptable excipients.

[0016] In a seventh aspect, there is provided an in vitro method of reducing the level of MEG3 expression in a cell, wherein the method comprises:

[0017] (a) introducing the oligonucleotide of the first aspect, the conjugate of the fourth aspect, or the pharmaceutical composition of the sixth aspect, into the cell, under suitable conditions for the oligonucleotide to bind to MEG3 transcripts, or

[0018] (b) introducing the virus particle of the third aspect, the delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, into the cell, under suitable conditions for the oligonucleotide comprised within the virus particle or delivery particle to bind to MEG3 transcripts, or (c) introducing the vector of the second aspect, or the pharmaceutical composition of the sixth aspect, into the cell, under suitable conditions for the vector to be transcribed within the cell to produce the oligonucleotide, and for the oligonucleotide to bind to MEG3 transcripts, or

[0019] (d) introducing the virus particle of the third aspect, the delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, into the cell, under suitable conditions for the vector comprised within the virus particle or delivery particle to be transcribed within the cell to produce the oligonucleotide, and for the oligonucleotide to bind to MEG3 transcripts.

[0020] In an eight aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use in research.

[0021] In an ninth aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use as a medicament.

[0022] In an tenth aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use in the treatment or prevention of a neurodegenerative disease, preferably Alzheimer’s disease or, spinal muscular atrophy, Parkinson's disease, Huntington’s disease, prion diseases such as Creutzfeldt-Jakob disease, motor neuron disease, amyotrophic lateral sclerosis, multiple sclerosis, frontotemporal dementia, Lewy body dementia, ataxia, spinocerebellar ataxia, multiple system atrophy, progressive supranuclear palsy, spinal muscular atrophy, spinal and bulbar muscular atrophy (SBMA) or Duchenne muscular dystrophy.

[0023] In an eleventh aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use in the treatment or prevention of a cardiovascular disease, preferably cardiac fibrosis.

[0024] In an twelfth aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use in the treatment or prevention of a lung disease, preferably obstructive pulmonary disease.

[0025] In an thirteenth aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use in the treatment or prevention of cancer retinopathy.

[0026] In an fourteenth aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use in the treatment or prevention of osteoarthritis.

[0027] In an fifteenth aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use in the treatment or prevention of diabetes.

[0028] In an sixteenth aspect, there is provided a method of treatment of a subject having a disease, or for preventing a disease in a subject, wherein the method comprises administering to the subject an effective amount of the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect.

[0029] In an seventeenth aspect, there is provided a method of inhibiting necroptosis in a subject in need thereof, wherein the method comprises administering to the subject an effective amount of the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect.

[0030] In an eighteenth aspect, there is provided the oligonucleotide of the first aspect, vector of the second aspect, virus particle of the third aspect, conjugate of the fourth aspect, delivery particle of the fifth aspect, or the pharmaceutical composition of the sixth aspect, for use in research.

[0031] DETAILED DESCRIPTION OF THE INVENTION

[0032] Features and embodiments further to the above aspects are described in further detail below. It should be noted that any feature, characteristic, embodiment or definition described herein is not limited to any particular aspect of the invention, and may be combined with any other aspect in any workable combination.

[0033] The inventors have discovered that certain sites on the MEG3 transcript, when targeted using oligonucleotides such as ASOs, lead to highly significant reductions in the level of MEG3 expression. For example, reductions in expression of over 90% and up to as much as 96% can be achieved. Unexpectedly, there is a certain region of the MEG3 transcript within which most effective knockdown is achieved. By performing an oligonucleotide ‘microwalk’ experiment, the inventors have confirmed that substantially the whole of this region provides for effective reductions in MEG3 expression when targeted using oligonucleotides such as ASOs. To the best of the inventors’ knowledge, it is highly unusual for there to be such hotspots of knockdown activity for any given RNA transcript.

[0034] Accordingly, the inventors have developed a set of oligonucleotides targeting these so-called ‘hotspot’ sites discovered in the MEG3 transcript that are capable of achieving significant reductions in the level of MEG3 expression. Advantageously, this provides a pool of validated oligonucleotides from which lead therapeutics may be developed for the treatment of diseases in which MEG3 is implicated such as neurodegenerative diseases (e.g. Alzheimer’s disease). A further advantage of the present discovery is that the oligonucleotides developed by the inventors target sites that are present in multiple isoforms of MEG3 transcript including the canonical isoform, and are target sites that are thought to be present in the majority of MEG3 transcripts present within a cell giving them broad applicability. It is believed that these attributes enhance the phenotype that is obtainable when using these oligonucleotides to target MEG3 expression. Additionally, the oligonucleotides display no cytotoxicity when transfected into cells, which is an important requirement for their development towards therapeutic use for example.

[0035] Definitions

[0036] As used herein, unless otherwise clear from context, the term “a” or “an” may be understood to mean “at least one”; the term “a,” “an” or “the” include plural reference unless the context clearly dictates otherwise; the term “or” may be understood to mean “and / or”; the terms “comprising”, “comprise”, “including” (whether used with “but not limited to” or not) and “include” (whether used with “but not limited to” or not) may be understood to encompass itemised components or steps whether presented by themselves or together with one or more additional components or steps; the term “another” may be understood to mean at least an additional / second one or more; and where ranges are provided, endpoints are included.

[0037] The terms “about” and “approximately” as used herein may be understood to permit standard variation as would be understood by those of ordinary skill in the art. In reference to a number, these terms are generally taken to include numbers that fall within a range of 5%, 10%, 15%, 20%, 25%, or 30%, in either direction (greater than or less than) of the number unless otherwise stated or otherwise evident from the context.

[0038] The terms “comprise” and “contain” mean “including but not limited to”, and are not intended to (and do not) exclude other features, integers, components, additives, or the like.

[0039] The term “nucleotide” as used herein refers to a molecule comprising a nucleobase, a sugar, and one or more internucleoside linkages. Nucleobases include naturally occurring bases guanine (G), adenine, (A), cytosine, (C), thymine, (T) and uracil (II) which are derivatives of purine or pyrimidine, as well as non-natural derivatives and modified or artificial analogues thereof. Sugars include the naturally occurring pentose (five-carbon sugar) deoxyribose (which forms DNA) and ribose (which forms RNA), as well as non-naturally occurring derivatives or analogues thereof such as morpholine rings. Nucleotides are linked through internucleoside linkages, which include phosphate groups (i.e. phosphodiester linkages) as well as several other derivatives or analogues thereof such as phosphorothioate groups, boranophosphate groups, and more. The term nucleotide encompasses any combination of nucleobases, modified nucleobases, sugars, modified sugars, phosphate bridges or modified internucleoside linkages. A natural nucleotide refers to a naturally occurring base, pentose sugar and phosphate internucleoside linkage. Unless otherwise specified, the prefix polyrefers to a nucleic acid containing 2 to about 10,000 nucleotide monomer units and the prefix oligo- refers to a nucleic acid containing 2 to about 200 nucleotide monomer units.

[0040] The term “nucleoside” as used herein refers to a nucleotide without the internucleoside linkage such as the phosphate group or equivalent thereof.

[0041] The term “nucleic acid” as used herein refers to nucleotides and polymers thereof.

[0042] The term “oligonucleotide” as used herein refers to a polymeric form of nucleotides of any length, typically up to about 200 nucleotides or base pairs, of ribonucleotides (RNA), deoxyribonucleotides (DNA), artificial nucleotides, or any combination thereof. Oligonucleotide may refer to an oligomer of nucleotides formed by linking several, for example up to 10, 20, 30, 40, 50, 60, 70, 80, 90, or up to 100 or more such as up to 200 contiguous nucleotides through covalent bonds between hydroxyl groups in the sugar moiety (or artificial equivalent thereof) and phosphate groups (or equivalents thereof). Inside the structure of an oligonucleotide, the phosphate group is generally considered to form an internucleoside bond or linkage. Oligonucleotides may be single-stranded or double-stranded. A single-stranded oligonucleotide can have double-stranded regions (formed by two single-stranded oligonucleotides annealed together through base pair complementarity) and a double- stranded oligonucleotide can have single-stranded regions, for example at regions where the two oligonucleotide chains are not complementary to each other.

[0043] The term “oligonucleotide” as used herein refers to the primary structure (i.e. sequence) of the molecules and thus includes for example double-stranded DNA, single-stranded DNA, partially double-stranded DNA, double-stranded RNA, single-stranded RNA, partially doublestranded RNA, double-stranded DNA-RNA hybrid molecules, single-stranded DNA-RNA hybrid molecules, and partially double-stranded DNA-RNA hybrid molecules, any one of which may comprise artificial nucleotides. Included are DNA and RNA analogues comprising modified nucleotides, modified sugar moieties, and modified internucleoside linkages. Nonlimiting examples of oligonucleotides include antisense oligonucleotides (ASOs), short interfering RNAs (siRNAs), short hairpin RNAs (shRNAs), anti-miRNAs (antimiRs), spliceswitching oligonucleotides and DNA or RNA aptamers.

[0044] “Ribonucleotide” may be abbreviated RNA; “deoxyribonucleotide” may be abbreviated DNA.

[0045] Unless otherwise specified, description of oligonucleotides and elements thereof are referred to from 5’ to 3’ as is conventional in the art.

[0046] Unless otherwise specified, oligonucleotides described herein may be provided and / or utilised in salt forms, particularly pharmaceutically acceptable salt forms. As those skilled in the art will appreciate after reading the present disclosure, in some embodiments, oligonucleotides may be provided as salts including but not limited to, e.g., sodium or potassium salt forms. As those skilled in the art will appreciate, in some embodiments, individual oligonucleotides within a composition may be considered to be of the same constitution and / or structure even though, within such composition (e.g., a liquid composition), particular such oligonucleotides might be in different salt form(s) (and may be dissolved and the oligonucleotide chain may exist as an anion form when, e.g., in a liquid composition) at a particular moment in time. For example, those skilled in the art will appreciate that, at a given pH, individual internucleoside linkages along an oligonucleotide chain may be in an acid (H) form, or in one of a plurality of possible salt forms (e.g., a sodium salt, or a salt of a different cation, depending on which ions might be present in the preparation or composition), and will understand that, so long as their acid forms (e.g., replacing all cations, if any, with H) are of the same constitution and / or structure, such individual oligonucleotides may properly be considered to be of the same constitution and / or structure.

[0047] The term “pharmaceutical composition” as used herein refers to an active agent, formulated together with one or more pharmaceutically acceptable carriers. In some embodiments, active agent is present in a unit dose amount appropriate for administration in a therapeutic regimen that shows a statistically significant probability of achieving a controlled therapeutic effect when administered to a relevant population.

[0048] The phrase “pharmaceutically acceptable” as used herein refers to those compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0049] The term “pharmaceutically acceptable salt” as used herein refers to salts of such compounds that are appropriate for use in pharmaceutical contexts, i.e. , salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge, et al. describes pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 66: 1-19 (1977). In some embodiments, a provided compound comprises more than one acid groups, for example, a provided oligonucleotide may comprise two or more acidic groups (e.g., in natural phosphate linkages and / or modified internucleoside linkages). In some embodiments, a pharmaceutically acceptable salt, or generally a salt, of such a compound comprises two or more cations, which can be the same or different. In some embodiments, in a pharmaceutically acceptable salt (or generally, a salt), each acidic group having sufficient acidity independently exists as its salt form (e.g., in an oligonucleotide comprising natural phosphate linkages and phosphorothioate internucleoside linkages, each of the natural phosphate linkages and phosphorothioate internucleoside linkages independently exists as its salt form). In some embodiments, a pharmaceutically acceptable salt of an oligonucleotide is a sodium salt of a provided oligonucleotide. In some embodiments, a pharmaceutically acceptable salt of an oligonucleotide is a sodium salt of a provided oligonucleotide, wherein each acidic linkage, e.g., each natural phosphate linkage and phosphorothioate internucleoside linkage, exists as a sodium salt form (all sodium salt).

[0050] The term “subject” or “test subject” as used herein refers to any organism to which a provided compound or composition is administered in accordance with the present disclosure e.g., for experimental, diagnostic, prophylactic, and / or therapeutic purposes. In some embodiments, a subject may be suffering from and / or susceptible to a disease, disorder, and / or condition. The term “substantially” as used herein refers to the qualitative condition of exhibiting total or near-total extent or degree of a characteristic or property of interest. One of ordinary skill in the biological arts will understand that biological and chemical phenomena rarely, if ever, go to completion and / or proceed to completeness or achieve or avoid an absolute result. The term “substantially” is therefore used herein to capture the potential lack of completeness inherent in many biological and / or chemical phenomena.

[0051] The phrase “suffering from” as used herein in relation to an individual who is suffering from a disease, disorder, and / or condition refers to an individual who has been diagnosed with and / or displays one or more symptoms of said disease, disorder, and / or condition.

[0052] The phrase “therapeutic agent” as used herein refers to any agent that, when administered to a subject, has a therapeutic effect and / or elicits a desired biological and / or pharmacological effect. In some embodiments, a therapeutic agent is any substance that can be used to alleviate, ameliorate, relieve, inhibit, prevent, delay onset of, reduce severity of, and / or reduce incidence of one or more symptoms or features of a disease, disorder, and / or condition.

[0053] The term “therapeutically effective amount” as used herein, in some embodiments, means an amount of a substance (e.g., a therapeutic agent, composition, and / or formulation) that elicits a desired biological response when administered as part of a therapeutic regimen. In some embodiments, a therapeutically effective amount of a substance is an amount that is sufficient, when administered to a subject suffering from or susceptible to a disease, disorder, and / or condition, to treat, diagnose, prevent, and / or delay the onset of the disease, disorder, and / or condition. As will be appreciated by those of ordinary skill in this art, the effective amount of a substance may vary depending on such factors as the desired biological endpoint, the substance to be delivered, the target cell or tissue, etc. For example, the effective amount of compound in a formulation to treat a disease, disorder, and / or condition is the amount that alleviates, ameliorates, relieves, inhibits, prevents, delays onset of, reduces severity of and / or reduces incidence of one or more symptoms or features of the disease, disorder, and / or condition. In some embodiments, a therapeutically effective amount is administered in a single dose; in some embodiments, multiple unit doses are required to deliver a therapeutically effective amount, in some embodiments, a single dose is an infusion, which may take up to one or more hours.

[0054] The term “treat,” “treatment,” or “treating” as used herein refers to any method used to partially or completely alleviate, ameliorate, relieve, inhibit, prevent, delay onset of, reduce severity of, and / or reduce incidence of one or more symptoms or features of a disease, disorder, and / or condition. T reatment may be administered to a subject who does not exhibit signs of a disease, disorder, and / or condition. In some embodiments, treatment may be administered to a subject who exhibits only early signs of the disease, disorder, and / or condition, for example for the purpose of decreasing the risk of developing pathology associated with the disease, disorder, and / or condition.

[0055] The term “identity” as used herein refers to the overall relatedness between polymeric molecules, e.g., between oligonucleotides. In some embodiments, polymeric molecules are considered to be “substantially identical” to one another if their sequences are at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% identical. Calculation of the percent identity of two nucleic acid or polypeptide sequences, for example, can be performed by aligning the two sequences for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second sequences for optimal alignment and non-identical sequences can be disregarded for comparison purposes). In certain embodiments, the length of a sequence aligned for comparison purposes is at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or substantially 100% of the length of a reference sequence. The nucleotides at corresponding positions are then compared. When a position in the first sequence is occupied by the same residue (e.g., nucleotide or amino acid) as the corresponding position in the second sequence, then the molecules are identical at that position. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which needs to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm. For example, the percent identity between two nucleotide sequences can be determined using the algorithm of Meyers and Miller (CABIOS, 1989, 4: 11-17), which has been incorporated into the ALIGN program (version 2.0). Nucleic acid sequence comparisons may be made with the ALIGN program using a PAM 120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. The percent identity between two nucleotide sequences can, alternatively, be determined using the GAP program in the GCG software package using an NWSgapdna.CMP matrix or with established pairwise alignment algorithms such as the Smith- Waterman or Needleman-Wunsch algorithm, or multiple sequence tools such as Clustal Omega or T-Coffee.

[0056] Throughout the description and claims of this specification, the singular encompasses the plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.

[0057] MEG3

[0058] The present invention provides an oligonucleotide for reducing the level of MEG3 expression in a cell, wherein the oligonucleotide is capable of binding to a target sequence within, suitably at least partially within, a region of the MEG3 transcript selected from the group consisting of: SEQ ID NO: 1 (Ensembl exon ID ENSE00002476600), SEQ ID NO: 2 (ENSE00001586203) and SEQ ID NO: 3 (ENSE00002440436). Any such oligonucleotides may be referred to herein as the oligonucleotide of the invention.

[0059] MEG3 (Maternally Expressed Gene 3) transcript is a long non-coding RNA (IncRNA) that is transcribed from the MEG3 gene. The MEG3 gene therefore expresses the MEG3 long noncoding RNA (IncRNA) transcript. In humans, the MEG3 gene is located on chromosome 14 (GRCh38, Genome Reference Consortium Human Build 38). MEG3 is strongly up-regulated in human neurons (Balusu et al. 2023).

[0060] Several variants of MEG3 transcript are documented for example under NCBI gene ID 55384, and under Ensembl gene ID ENSG00000214548. These transcript variants or isoforms each contain a different combination of exons due to alternative splicing.

[0061] A canonical transcript is defined by Ensemble as the transcript that, on balance, has the highest coverage of conserved exons, highest expression, longest coding sequence and is represented in other key resources, such as NCBI and UniProt. A canonical transcript typically represents the most biologically significant transcript produced from a given gene.

[0062] Exons ENSE00002476600 (SEQ ID NO: 1), ENSE00001586203 (SEQ ID NO: 2) and ENSE00002440436 (SEQ ID NO: 3), are comprised within the canonical transcript of MEG3 according to Ensembl (transcript MEG-266; Ensembl transcript ID ENST00000556407.5). The length of these three exons are 371 , 109 and 152 nucleotides (nt), respectively. The length of the fully spliced (i.e. mature) MEG-226 transcript is 632 nt. Therefore, advantageously, the exons targeted by the oligonucleotides of the invention are present in most isoforms of the MEG3 transcript.

[0063] Suitably therefore the oligonucleotide of the invention is capable of binding to a target sequence within, or at least partially within, a region of the canonical MEG3 transcript. Suitably wherein the canonical MEG3 transcript is defined according to Ensembl as transcript MEG- 266; Ensembl transcript ID ENST00000556407.5. By “partially within” it is meant that a part of the target sequence may be located within any region of the MEG3 transcript as defined herein, and a part of the target sequence may be located outside of any region of the MEG3 transcript as defined herein. Suitably such that the target sequence may overlap a border, such as the 5’ or 3’ border, of any one or more regions of the MEG3 transcript as defined herein, suitably the target sequence may overlap the 5’ or 3’ border of any one or more of SEQ I D NO: 1 , 2 or 3, suitably the target sequence may overlap the 5’ or 3’ border of any other SEQ ID NO mentioned herein. For example, the target sequence may overlap the 3’ border of a first region of the MEG3 transcript as defined herein, and the 5’ region of a downstream, second region of the MEG3 transcript as defined herein. Suitably, ‘partially within’ may mean that at least 50% of a target sequence is within a region of MEG3 transcript as defined elsewhere herein, suitably at least 5, 6, 7, 8, 9 or at least 10 nucleotides are within a region of the MEG3 transcript as defined elsewhere herein, suitably at least 5, 6, 7, 8, 9 or at least 10 nucleotides are within a region of MEG3 transcript selected from any of SEQ ID NO:1 , 2 or 3. Suitably however, in preferred embodiments, the target sequence is located within, suitably wholly within, any region of the MEG3 transcript as defined herein, suitably wholly within SEQ ID NO:1 , 2 or 3, suitably wholly within any one or more of. By “located wholly within” is meant that the whole of the target sequence, suitably the entire target sequence, is within a region of the MEG3 transcript as defined hereinabove, suitably within one of SEQ I D NO: 1 , 2 or 3.

[0064] Region

[0065] The oligonucleotides of the invention are capable of binding to a target sequence which is located within, suitably at least partially within, a region of the MEG3 transcript. Suitably the region of the MEG3 transcript in which the target sequence is located or at least partially located is defined below.

[0066] It will be understood that, an oligonucleotide which is capable of binding to a target sequence within, or at least partially within, a region of the MEG3 transcript will typically also be capable of binding to the same region in the MEG3 gene. The “region” as used herein is defined by sequence. Suitably, the oligonucleotide may be capable of binding to a target sequence within, or at least partially within, a region of the MEG3 gene. It is understood in the art that a transcript is single-stranded and that a gene is double stranded. Accordingly, if an oligonucleotide is capable of binding to a region of MEG3 transcript, that oligonucleotide as well as the reverse complement of that oligonucleotide will typically both be capable of binding to that same region in the MEG3 gene. As used herein, the “region” of the MEG3 transcript refers to a stretch or length of nucleotides comprised in the sequence of the MEG3 transcript, which contains or at least partially contains the target sequence to which the oligonucleotide is capable of binding. In a preferred embodiment, the region contains the whole of i.e. the entirety of the target sequence to which the oligonucleotide is capable of binding.

[0067] Suitably the region of the MEG3 transcript will physically exist in unspliced MEG3 transcripts, in spliced MEG3 transcripts (partially or fully spliced transcripts), and in the MEG3 gene. The oligonucleotide may therefore be capable of binding to unspliced MEG3 transcripts, spliced MEG3 transcripts, and / or the MEG3 gene. Suitably therefore, the region (which at least partially comprises the target sequence to which the oligonucleotide is capable of binding) may be comprised in an unspliced MEG3 transcript, a spliced MEG3 transcript and / or the MEG3 gene. An “unspliced” transcript refers to a transcript containing introns; a “spliced” transcript refers to a transcript from which some introns (partially spliced transcripts) or all introns (fully spliced transcripts) have been spliced out.

[0068] Suitably the region may comprise or consist of one or more exons or exonic sequences of the MEG3 transcript. Suitably, the region may comprise or consist of one or more canonical exons or canonical exonic sequences of the MEG3 transcript. Suitably, the region may comprise or consist of one or more exons or exonic sequences of the MEG3 gene. Suitably, the region may comprise or consist of one or more canonical exons or canonical exonic sequences of MEG3 gene.

[0069] The region may comprise or consist of any one or more of: SEQ ID NO: 1 (ENSE00002476600), SEQ ID NO: 2 (ENSE00001586203), and / or SEQ ID NO: 3 (ENSE00002440436). Suitably, the region may comprise or consist of SEQ ID NO: 1

[0070] (ENSE00002476600). Suitably, the region may comprise or consist of SEQ ID NO: 2

[0071] (ENSE00001586203). Suitably, the region may comprise or consist of SEQ ID NO: 3

[0072] (ENSE00002440436).

[0073] In one embodiment, the region is selected from the group consisting of: SEQ ID NO: 1 (ENSE00002476600), SEQ ID NO: 2 (ENSE00001586203), and SEQ ID NO: 3 (ENSE00002440436). In one embodiment, the region is selected from the group consisting of: SEQ ID NO: 1 and SEQ ID NO: 2. In one embodiment, the region is selected from the group consisting of: SEQ ID NO: 1 and SEQ ID NO: 3. In one embodiment, the region is selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 3. As demonstrated in the Examples using ASO12 and ASO13, all oligonucleotides tested by the inventors that are capable of binding to a target sequence within the 3’ region of SEQ ID NO: 2 (ENSE00001586203) are particularly effective at reducing the level of MEG3 expression. Said 3’ region of ENSE00001586203 is designated SEQ ID NO: 4 and represents a notable hotspot of knockdown activity when targeted using oligonucleotides. Suitably therefore, the region may comprise or consist of SEQ ID NO: 4 (the 3’ region of ENSE00001586203 downstream of and including the exemplified ASO12 target site). In one embodiment, the region is selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 3.

[0074] As demonstrated in the Examples, substantially all oligonucleotides tested by the inventors that are capable of binding to SEQ ID NO: 3 (ENSE00002440436) unexpectedly lead to significant reductions in the level of MEG3 expression. Oligonucleotides capable of binding SEQ ID NO: 3 virtually abolish all MEG3 expression. SEQ ID NO: 3 (ENSE00002440436) therefore represents a notable hotspot of knockdown activity when targeted using oligonucleotides. In one embodiment therefore, the region is SEQ ID NO: 3 (ENSE00002440436).

[0075] Subregions of ENSE00002440436 (SEQ ID NO: 3)

[0076] The 5’ region of ENSE00002440436, i.e. substantially the whole of ENSE00002440436 upstream of the exemplified ASO19 target site, represents an even more notable hotspot of knockdown activity when targeted using oligonucleotides. Said 5’ region of ENSE00002440436 is designated as SEQ ID NO: 5. Suitably therefore, the region may comprise or consist of SEQ ID NO: 5 (the 5’ region of ENSE00002440436 upstream of the exemplified ASO19 target site). Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 5. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 5. In one embodiment, the region is selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 5. In one embodiment, the region is SEQ ID NO: 5.

[0077] With reference to the Examples, a subregion of SEQ ID NO: 5, namely the 5’ region of ENSE00002440436 capable of being bound by exemplified oligonucleotides ASO14-16 and ASO20-31 (said region being designated SEQ ID NO: 6), is also a notable hotspot of knockdown activity when targeted using oligonucleotides. Suitably therefore, the region may comprise or consist of SEQ ID NO: 6 (the 5’ region of ENSE00002440436 upstream of and including the exemplified oligonucleotide ASO16 target site). Suitably, in addition, the exemplified ASO18 target site, SEQ ID NO: 105, is also shown to be an effective region. Suitably, therefore, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 6 and SEQ ID NO: 105. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 6. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2, SEQ ID NO: 6 and SEQ ID NO: 105. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 6. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 4, SEQ ID NO: 6, SEQ ID NO: 105. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 6. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 6 and SEQ ID NO: 105. In one embodiment, the region is SEQ ID NO: 6.

[0078] Subregions of SEQ ID NO: 6

[0079] Further subregions of SEQ ID NO: 6 are yet more notable hotspots of knockdown activity when targeted using oligonucleotides of the invention. These subregions include the 5’ region capable of being bound by exemplified oligonucleotides ASO14-15 and ASO20-23 (said region being designated SEQ ID NO: 7). Suitably therefore, the region may comprise or consist of SEQ ID NO: 7 (the 5’ region of ENSE00002440436 upstream of and including the exemplified ASO15 target site). Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 7. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 7. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 7. Suitably, the region may be SEQ ID NO: 7.

[0080] These further subregions also include the region capable of being bound by exemplified oligonucleotides ASO26-28 (said region being designated SEQ ID NO: 8). Suitably therefore, the region may comprise or consist of SEQ ID NO: 8 (a central region of ENSE00002440436 including the exemplified ASO26-28 target sites). Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 8. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 8. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 8. Suitably, the region may be SEQ ID NO: 8.

[0081] These further subregions also include the region capable of being bound by exemplified oligonucleotides ASO30, 31 and 16 (said region being designated SEQ ID NO: 9). Suitably therefore, the region may comprise or consist of SEQ ID NO: 9 (a central region of ENSE00002440436 including the exemplified ASO30, 31 and 16 target sites). Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 9. Suitably, the region may be SEQ ID NO: 9.

[0082] Suitably, the region may be any combination of the above mentioned subregions of SEQ ID NO: 3 (ENSE00002440436) that were notable hotspots of knockdown activity when targeted using oligonucleotides. Suitably therefore, the region may be selected from the group consisting of: SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 7 and SEQ ID NO: 8. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 7 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 8 and SEQ ID NO: 9. Any of these groups may further consist of: SEQ ID NO: 105, e.g. the region may be selected from the group consisting of: SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and SEQ ID NO: 105.

[0083] Further definitions of the region

[0084] Suitably, the combinations of subregions of SEQ ID NO: 3 may be comprised in a group of regions further comprising SEQ ID NO: 1 and SEQ ID NO: 2. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 7 and SEQ ID NO: 8. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 7 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 8 and SEQ ID NO: 9. Any of these groups may further consist of: SEQ ID NO: 105, e.g. the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and SEQ ID NO: 105.

[0085] Suitably, the combinations of subregions of SEQ ID NO: 3 may be comprised in a group of regions further comprising SEQ ID NO: 1 and SEQ ID NO: 4. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 4, SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 4, SEQ ID NO: 7 and SEQ ID NO: 8. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 4, SEQ ID NO: 7 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1, SEQ ID NO: 4, SEQ ID NO: 8 and SEQ ID NO: 9. Any of these groups may further consist of: SEQ ID NO: 105, e.g. the region may be selected from the group consisting of: SEQ ID NO:

[0086] 1 , SEQ ID NO: 4, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and SEQ ID NO: 105.

[0087] Suitably, the combinations of subregions of SEQ ID NO: 3 may be comprised in a group of regions further comprising SEQ ID NO: 2. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2, SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2, SEQ ID NO: 7 and SEQ ID NO: 8. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2, SEQ ID NO: 7 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2, SEQ ID NO: 8 and SEQ ID NO: 9. Any of these groups may further consist of: SEQ ID NO: 105, e.g. the region may be selected from the group consisting of: SEQ ID NO: 2, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and SEQ ID NO: 105.

[0088] Suitably, the combinations of subregions of SEQ ID NO: 3 may be comprised in a group of regions further comprising with SEQ ID NO: 4. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 4, SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 4, SEQ ID NO: 7 and SEQ ID NO: 8. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 4, SEQ ID NO: 7 and SEQ ID NO: 9. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 4, SEQ ID NO: 8 and SEQ ID NO: 9. Any of these groups may further consist of: SEQ ID NO: 105, e.g. the region may be selected from the group consisting of: SEQ ID NO: 4, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and SEQ ID NO: 105.

[0089] Suitably, the region may encompass both SEQ ID NO: 7 and SEQ ID NO: 8, said region being designated SEQ ID NO: 119. Suitably therefore, the region may comprise or consist of SEQ ID NO: 119, suitably in addition to the exemplified ASO18 target site, SEQ ID NO: 105. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 119 and SEQ ID NO: 105. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 119. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2, SEQ ID NO: 119 and SEQ ID NO: 105. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 119. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 4, SEQ ID NO: 119, SEQ ID NO: 105. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 119. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 119 and SEQ ID NO: 105. In one embodiment, the region is SEQ ID NO: 119. Suitably, the region may encompass both SEQ ID NO: 8 and SEQ ID NO: 9, said region being designated SEQ ID NO: 120. Suitably therefore, the region may comprise or consist of SEQ ID NO: 120, suitably in addition to the exemplified ASO18 target site, SEQ ID NO: 105. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 120 and SEQ ID NO: 105. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 120. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2, SEQ ID NO: 120 and SEQ ID NO: 105. Suitably, the region may be selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 120. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 4, SEQ ID NO: 120, SEQ ID NO: 105. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 120. In one embodiment, the region may be selected from the group consisting of: SEQ ID NO: 120 and SEQ ID NO: 105. In one embodiment, the region is SEQ ID NO: 120.

[0090] As mentioned hereinabove, the region may be comprised in a spliced MEG3 transcript. As such, it is envisioned that the oligonucleotide of the invention may be capable of binding across exon-intron of unspliced MEG3 transcripts and / or exon-exon sites of spliced MEG3 transcripts. Suitably, the region may comprise or consist of any spliced MEG3 transcripts represented by any combination of SEQ ID NO: 1 , SEQ ID NO: 2 or SEQ ID NO: 3 (including appropriate exon-containing subregions thereof as defined herein by SEQ ID NO: 4-7). Accordingly, the region may comprise or consist of a single continuous sequence that is a direct combination of SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 3 (SEQ ID NO: 48), a direct combination of SEQ ID NO: 2 and SEQ ID NO: 3 (SEQ ID NO: 49), a direct combination of SEQ ID NO: 4 and SEQ ID NO: 3 (SEQ ID NO: 50), a direct combination of SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 5 (SEQ ID NO: 121), a direct combination of SEQ ID NO: 2 and SEQ ID NO: 5 (SEQ ID NO: 51), a direct combination of SEQ ID NO: 4 and SEQ ID NO: 5 (SEQ ID NO: 52), a direct combination of SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 6 (SEQ ID NO: 122), a direct combination of SEQ ID NO: 2 and SEQ ID NO: 6 (SEQ ID NO: 53), a direct combination of SEQ ID NO: 4 and SEQ ID NO: 6 (SEQ ID NO: 54), a direct combination of SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 7 (SEQ ID NO: 123), a direct combination of SEQ ID NO: 2 and SEQ ID NO: 7 (SEQ ID NO: 55), a direct combination of SEQ ID NO: 4 and SEQ ID NO: 7 (SEQ ID NO: 56), a direct combination of SEQ ID NO: 1 , SEQ ID NO: 2 and SEQ ID NO: 119 (SEQ ID NO: 124), a direct combination of SEQ ID NO: 2 and SEQ ID NO: 119 (SEQ ID NO: 125), a direct combination of SEQ ID NO: 4 and SEQ ID NO: 119 (SEQ ID NO: 126). Suitably, the region may comprise or consist of any one of SEQ ID NO: 48-56, 121- 126 (these being representative of regions of spliced MEG3 transcripts). In one preferred embodiment, the region is SEQ ID NO: 3.

[0091] The target sequence may be comprised within any one or more of SEQ ID NO: 1-9, 48-56, 119-126 (i.e. any one or more of SEQ ID NO: 1 , 2, 3, 4, 5, 6, 7, 8, 9, 48, 49, 50, 51 , 52, 53, 54, 55, 56, 119, 120, 121 , 122, 123, 124, 125 and / or 126).

[0092] The region may be one which, when targeted using an oligonucleotide, leads to a certain amount reduction in the level of MEG3 expression, as described in more detail hereinbelow.

[0093] Target sequence

[0094] The target sequence as referred to herein is the specific nucleic acid sequence located within or at least partially (such as fully) within at least one of the regions recited herein that is bound by the oligonucleotide of the invention. Therefore the target sequence may be referred to as a target nucleic acid sequence.

[0095] Suitably the target sequence may be located in an unspliced MEG3 transcript, a spliced MEG3 transcript, and / or the MEG3 gene.

[0096] Suitably the target sequence may be comprised at least partially (such as wholly) within one or more exons or exonic sequences of MEG3 transcript. Suitably, the target sequence may be comprised at least partially (such as wholly) within one or more canonical exons or canonical exonic sequences of MEG3 transcript. Suitably, the target sequence may be comprised at least partially (such as wholly) within one or more exons or exonic sequences of the MEG3 gene. Suitably, the target sequence may be comprised at least partially (such as wholly) within one or more canonical exons or canonical exonic sequences of MEG3 gene.

[0097] Suitably the target sequence may be comprised within or at least partially within any region of MEG3 as defined hereinabove. For example, the target sequence may be comprised within or at least partially within any one or more of SEQ ID NO: 1-9, 48-56, 119-126.

[0098] Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 88- 118.

[0099] The target sequence may be comprised within SEQ ID NO: 1 (ENSE00002476600), suitably partially or wholly comprised within SEQ ID NO: 1. Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 88-94. Suitably, the target sequence may comprise or consist of SEQ ID NO: 88 and / or SEQ ID NO: 92. Suitably the target sequence may comprise or consist of SEQ ID NO: 88. Suitably, the target sequence may comprise or consist of SEQ ID NO: 92.

[0100] The target sequence may be comprised within SEQ ID NO: 2 (ENSE00001586203) , suitably partially or wholly comprised within SEQ ID NO: 2. Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 95-100. Suitably, the target sequence may comprise or consist of SEQ ID NO: 95-100. Suitably, the target sequence may comprise or consist of SEQ ID NO: 95, 99 and / or 100. Suitably, the target sequence may comprise or consist of SEQ ID NO: 98-100. Suitably, the target sequence may comprise or consist of SEQ ID NO: 99 and / or 100. Suitably, the target sequence may comprise or consist of SEQ ID NO: 98. Suitably, the target sequence may comprise or consist of SEQ ID NO: 99. Suitably, the target sequence may comprise or consist of SEQ ID NO: 100.

[0101] The target sequence may be comprised within SEQ ID NO: 4 (the 3’ region of ENSE00001586203 downstream of and including the exemplified oligonucleotide ASO12 target site), suitably partially or wholly comprised within SEQ ID NO: 4. Suitably, the target sequence may comprise or consist of SEQ ID NO: 99 and / or SEQ ID NO: 100. Suitably, the target sequence may comprise or consist of SEQ ID NO: 99. Suitably, the target sequence may comprise or consist of SEQ ID NO: 100.

[0102] The target sequence may be comprised within SEQ ID NO: 3 (ENSE00002440436) , suitably partially or wholly comprised within SEQ ID NO: 3. Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 101-118. Suitably, the target sequence may comprise or consist of any one or more of those target sequences that are comprised within any one or more of SEQ ID NO: 5-9 as mentioned hereinbelow (since SEQ ID NO: 5-9 are comprised within SEQ ID NO: 3).

[0103] The target sequence may be comprised within SEQ ID NO: 5 (the 5’ region of ENSE00002440436 upstream of the exemplified oligonucleotide ASO19 target site) , suitably partially or wholly comprised within SEQ ID NO: 5. Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 101-105, 107-118. Suitably, the target sequence may comprise or consist of any one or more of those target sequences that are comprised within any one or more of SEQ ID NO: 6-9 as mentioned hereinbelow (since SEQ ID NO: 6-9 are comprised within SEQ ID NO: 5).

[0104] The target sequence may be comprised within SEQ ID NO: 6 (the 5’ region of ENSE00002440436 upstream of and including the exemplified oligonucleotide ASO16 target site) , suitably partially or wholly comprised within SEQ ID NO: 6. Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 101-103, 107-118. Suitably, the target sequence may comprise or consist of any one or more of those target sequences that are comprised within any one or more of SEQ ID NO: 7-9 (since SEQ ID NO: 7-9 are comprised within SEQ ID NO: 6), suitably in combination with any one or more of SEQ ID NO: 111 , 112, 116.

[0105] The target sequence may be comprised within SEQ ID NO: 7 (the 5’ region of ENSE00002440436 upstream of and including the exemplified oligonucleotide ASO15 target site) , suitably partially or wholly comprised within SEQ ID NO: 7. Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 101 , 102, 107-110.

[0106] The target sequence may be comprised within SEQ ID NO: 8 (a central region of ENSE00002440436 including the target sites of exemplified oligonucleotides ASO26-28), suitably partially or wholly comprised within SEQ ID NO: 8. Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 113-115.

[0107] The target sequence may be comprised within SEQ ID NO: 9 (a central region of ENSE00002440436 including the target sites of exemplified oligonucleotide ASQ30, 31 , 16) , suitably partially or wholly comprised within SEQ ID NO: 9. Suitably, the target sequence may comprise or consist of any one or more of SEQ ID NO: 103, 117, 118.

[0108] The target sequence may equally be comprised within any one or more of the regions of the MEG3 transcript according to SEQ ID NO: 119, 120, 48-56, 121-126.

[0109] Suitably, the target sequence is comprised within SEQ ID NO: 119, suitably partially or wholly comprised within SEQ ID NO: 119, and may comprise target sequences comprised within SEQ ID NO: 7 and 8.

[0110] Suitably, the target sequence is comprised within SEQ ID NO: 120, suitably partially or wholly comprised within SEQ ID NO: 120, and may comprise target sequences comprised within SEQ ID NO: 8 and 9.

[0111] Suitably, the target sequence is comprised within SEQ ID NO: 48, suitably partially or wholly comprised within SEQ ID NO: 48, and may comprise target sequences comprised within SEQ ID NO: 1-3.

[0112] Suitably, the target sequence is comprised within SEQ ID NO: 49, suitably partially or wholly comprised within SEQ ID NO: 49, and may comprise target sequences comprised within SEQ ID NO: 2 and 3. Suitably, the target sequence is comprised within SEQ ID NO: 50, suitably partially or wholly comprised within SEQ ID NO: 50, and may comprise target sequences comprised within SEQ ID NO: 4 and 3.

[0113] Suitably, the target sequence is comprised within SEQ ID NO: 51 , suitably partially or wholly comprised within SEQ ID NO: 51 , and may comprise target sequences comprised within SEQ ID NO: 2 and 5.

[0114] Suitably, the target sequence is comprised within SEQ ID NO: 52, suitably partially or wholly comprised within SEQ ID NO: 52, and may comprise target sequences comprised within SEQ ID NO: 4 and 5.

[0115] Suitably, the target sequence is comprised within SEQ ID NO: 53, suitably partially or wholly comprised within SEQ ID NO: 53, and may comprise target sequences comprised within SEQ ID NO: 2 and 6.

[0116] Suitably, the target sequence is comprised within SEQ ID NO: 54, suitably partially or wholly comprised within SEQ ID NO: 54, and may comprise target sequences comprised within SEQ ID NO: 4 and 6.

[0117] Suitably, the target sequence is comprised within SEQ ID NO: 55, suitably partially or wholly comprised within SEQ ID NO: 55, and may comprise target sequences comprised within SEQ ID NO: 2 and 7.

[0118] Suitably, the target sequence is comprised within SEQ ID NO: 56, suitably partially or wholly comprised within SEQ ID NO: 56, and may comprise target sequences comprised within SEQ ID NO: 4 and 7.

[0119] Suitably, the target sequence is comprised within SEQ ID NO: 121 , suitably partially or wholly comprised within SEQ ID NO: 121 , and may comprise target sequences comprised within SEQ ID NO: 1 , 2 and 5.

[0120] Suitably, the target sequence is comprised within SEQ ID NO: 122, suitably partially or wholly comprised within SEQ ID NO: 122, and may comprise target sequences comprised within SEQ ID NO: 1 , 2 and 6.

[0121] Suitably, the target sequence is comprised within SEQ ID NO: 123, suitably partially or wholly comprised within SEQ ID NO: 123, and may comprise target sequences comprised within SEQ ID NO: 1 , 2 and 7. Suitably, the target sequence is comprised within SEQ ID NO: 124, suitably partially or wholly comprised within SEQ ID NO: 124, and may comprise target sequences comprised within SEQ ID NO: 1 , 2 and 119.

[0122] Suitably, the target sequence is comprised within SEQ ID NO: 125, suitably partially or wholly comprised within SEQ ID NO: 125, and may comprise target sequences comprised within SEQ ID NO: 2 and 119.

[0123] Suitably, the target sequence is comprised within SEQ ID NO: 126, suitably partially or wholly comprised within SEQ ID NO: 126, and may comprise target sequences comprised within SEQ ID NO: 4 and 119.

[0124] The target sequence may be one which, when targeted using an oligonucleotide, leads to a certain reduction in the level of MEG3 expression, as described in more detail hereinbelow.

[0125] The target sequence may be at least 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 35 or 40 or more nucleotides (or base pairs) in length. The target sequence may be up to 120, 110, 100, 90, 80, 70, 60, 50, 40, 30 or 20 nucleotides (or base pairs) or less in length. The target sequence may be anywhere between and including 5-40 nucleotides (or base pairs) in length. The target sequence may anywhere between and including 10-30 nucleotides (or base pairs) in length. In some embodiments, the target sequence is 10 nucleotides (or base pairs) in length. In some embodiments, the target sequence is 20 nucleotides (or base pairs) in length. In some embodiments, the target sequence is 30 nucleotides (or base pairs) in length.

[0126] Oligonucleotide length

[0127] Oligonucleotides of the present disclosure can be of various lengths. As used herein, the length of an oligonucleotide is quantified as the number of nucleotides in the longest single strand of that oligonucleotide unless the context indicates otherwise. For a single-stranded oligonucleotide, length refers to the length of that strand. For a completely double-stranded oligonucleotide, length refers to the length of either strand. For a partially double-stranded oligonucleotide, length refers to the longest single strand of that oligonucleotide.

[0128] The length of the oligonucleotide may vary according to the target sequence details of which are provided elsewhere hereinabove. Suitably the oligonucleotide may have about the same length as the target sequence (e.g. for an ASO or a siRNA) or about double the length of the target sequence (e.g. for a shRNA). Suitably, the oligonucleotide may comprise a length that is suitable to allow for binding to a target sequence as described elsewhere herein. The oligonucleotide such as an ASO, siRNA or shRNA may be any length required to allow binding of the oligonucleotide to the target sequence. Suitably to allow said binding under physiological conditions.

[0129] The oligonucleotide may be at least 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 35 or 40 or more nucleotides (or base pairs) in length. The oligonucleotide may be up to 120,

[0130] 110, 100, 90, 80, 70, 60, 50, 40, 30, 25, 23 20 or 15 nucleotides (or base pairs) or less in length.

[0131] Length of ASOs and siRNAs

[0132] Antisense oligonucleotides (ASOs) are typically 10-30 nucleotides in length.

[0133] Short interfering RNAs (siRNAs) are typically 10-30 base pairs in length, suitably 15-25 base pairs in length, suitably 18-23 base pairs in length.

[0134] Suitably, in some embodiments, the oligonucleotide may be at least 5 nucleotides (or base pairs) in length. In some embodiments, the oligonucleotide may be at least 10 nucleotides (or base pairs) in length. In some embodiments, the oligonucleotide may be at least 15 nucleotides (or base pairs) in length. In some embodiments, the oligonucleotide may be at least 20 nucleotides (or base pairs) in length.

[0135] Suitably, in some embodiments, the oligonucleotide may be no more than 30 nucleotides (or base pairs) in length. In some embodiments, the oligonucleotide may be no more than 25 nucleotides (or base pairs) in length. In some embodiments, the oligonucleotide may be no more than 20 nucleotides (or base pairs) in length.

[0136] Suitably, in some embodiments, the oligonucleotide may be 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39,

[0137] 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49, 50, 51 , 52, 53, 54, 55, 56, 57, 58, 59, 60, 61 , 62, 63, 64,

[0138] 65, 66, 67, 68, 69, 70, 71 , 72, 73, 74, 75, 76, 77, 78, 79, 80, 81 , 82, 83, 84, 85, 86, 87, 88, 89,

[0139] 90, 91 , 92, 93, 94, 95, 96, 97, 98, 99, 100, 101 , 102, 103, 104, 105, 106, 107, 108, 109, 110,

[0140] 111 , 112, 113, 114, 115, 116, 117, 118, 119 or 120 nucleotides (or base pairs) in length.

[0141] Suitably, in some embodiments, the oligonucleotide may be between 5-10, 5-11 , 5-12, 5-13, 5-14, 5-15, 5-16, 5-17, 5-18, 5-19, 5-20, 5-21 , 5-22, 5-23, 5-24, 5-25, 5-26, 5-27, 5-28, 5-29 or 5-30 nucleotides (or base pairs) in length. Suitably, in some embodiments, the oligonucleotide may be between 7-10, 7-11 , 7-12, 7-13, 7-14, 7-15, 7-16, 7-17, 7-18, 7-19, 7-20, 7-21 , 7-22, 7-23, 7-24, 7-25, 7-26, 7-27, 7-28, 7-29 or 7-30 nucleotides (or base pairs) in length.

[0142] Suitably, in some embodiments, the oligonucleotide may be between 10-11 , 10-12, 10-13, IQ- 14, 10-15, 10-16, 10-17, 10-18, 10-19, 10-20, 10-21 , 10-22, 10-23, 10-24, 10-25, 10-26, IQ- 27, 10-28, 10-29 or 10-30 nucleotides (or base pairs) in length.

[0143] Suitably, in some embodiments, the oligonucleotide may be between 12-13, 12-14, 12-15, 12- 16, 12-17, 12-18, 12-19, 12-20, 12-21 , 12-22, 12-23, 12-24, 12-25, 12-26, 12-27, 12-28, 12- 29 or 12-30 nucleotides (or base pairs) in length.

[0144] Suitably, in some embodiments, the oligonucleotide may be between 15-16, 15-17, 15-18, 15- 19, 15-20, 15-21 , 15-22, 15-23, 15-24, 15-25, 15-26, 15-27, 15-28, 15-29 or 15-30 nucleotides (or base pairs) in length.

[0145] Suitably, in some embodiments, the oligonucleotide may be between 17-18, 17-19, 17-20, 17-

[0146] 21 , 17-22, 17-23, 17-24, 17-25, 17-26, 17-27, 17-28, 17-29 or 17-30 nucleotides (or base pairs) in length.

[0147] Suitably, in some embodiments, the oligonucleotide may be between 18-19, 18-20, 18-21 , 18-

[0148] 22, 18-23, 18-24, 18-25, 18-26, 18-27, 18-28, 18-29 or 18-30 nucleotides (or base pairs) in length.

[0149] Suitably, in some embodiments, the oligonucleotide may be between 19-20, 19-21 , 19-22, 19-

[0150] 23, 19-24, 19-25, 19-26, 19-27, 19-28, 19-29 or 19-30 nucleotides (or base pairs) in length.

[0151] Suitably, in some embodiments, the oligonucleotide may be between 20-21 , 20-22, 20-23, 20-

[0152] 24, 20-25, 20-26, 20-27, 20-28, 20-29 or 20-30 nucleotides (or base pairs) in length.

[0153] In some embodiments, the oligonucleotide is between 10 to 30 nucleotides (or base pairs) in length. In some embodiments, the oligonucleotide is between 15 to 25 nucleotides (or base pairs) in length. In some embodiments, the oligonucleotide is between 18 to 23 nucleotides (or base pairs) in length.

[0154] In some embodiments, the oligonucleotide is an ASO and is 15 nucleotides in length. In one embodiment, the oligonucleotide is an ASO and is 20 nucleotides in length. In one embodiment, the oligonucleotide is an ASO and is 25 nucleotides in length. In one embodiment, the oligonucleotide is an ASO that is 30 nucleotides in length. In some embodiments, the oligonucleotide is an siRNA and is 15 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 18-23 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 18 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 19 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 20 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 21 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 22 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 23 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 24 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 25 base pairs in length. In one embodiment, the oligonucleotide is an siRNA and is 30 base pairs in length.

[0155] In some embodiments, the oligonucleotide is an siRNA comprising two strands of different lengths. Suitably, siRNAs are typically referred to as comprising a guide strand and a passenger strand. Suitably, the oligonucleotide may be an siRNA comprising a guide strand and a passenger strand, suitably of different lengths. Suitably, the passenger strand may be at least 15 nucleotides in length. Suitably, the guide strand may be up to 25 nucleotides in length.

[0156] Length of shRNAs

[0157] Short hairpin RNAs (shRNAs) are typically 40-70 nucleotides in length.

[0158] Suitably, in some embodiments, the oligonucleotide may be at least 30 base pairs (or nucleotides) in length. In some embodiments, the oligonucleotide may be at least 35 base pairs (or nucleotides) in length. In some embodiments, the oligonucleotide may be at least 40 base pairs (or nucleotides) in length.

[0159] Suitably, in some embodiments, the oligonucleotide may be no more than 120 base pairs (or nucleotides) in length. In some embodiments, the oligonucleotide may be no more than 100 base pairs (or nucleotides) in length. In some embodiments, the oligonucleotide may be no more than 80 base pairs (or nucleotides) in length. In some embodiments, the oligonucleotide may be no more than 70 base pairs (or nucleotides) in length. In some embodiments, the oligonucleotide may be no more than 60 base pairs (or nucleotides) in length. In some embodiments, the oligonucleotide may be no more than 50 base pairs (or nucleotides) in length. Suitably, in some embodiments, the oligonucleotide may be between 35-40, 35-41 , 35-42, 35- 43, 35-44, 35-45, 35-46, 35-47, 35-48, 35-49, 35-50, 35-51 , 35-52, 35-53, 35-54, 35-55, 35- 56, 35-57, 35-58, 35-59, 35-60, 35-61 , 35-62, 35-63, 35-64, 35-65, 35-66, 35-67, 35-68, 35-

[0160] 69 or 35-70 nucleotides (or base pairs) in length.

[0161] Suitably, in some embodiments, the oligonucleotide may be between 37-40, 37-41 , 35-42, 35-

[0162] 43, 35-44, 35-45, 35-46, 35-47, 35-48, 35-49, 35-50, 35-51 , 35-52, 35-53, 35-54, 35-55, 35-

[0163] 56, 35-57, 35-58, 35-59, 35-60, 35-61 , 35-62, 35-63, 35-64, 35-65, 35-66, 35-67, 35-68, 35- 69, 35-70 nucleotides (or base pairs) in length.

[0164] Suitably, in some embodiments, the oligonucleotide may be between 40-41 , 40-42, 40-43, 40-

[0165] 44, 40-45, 40-46, 40-47, 40-48, 40-49, 40-50, 40-51 , 40-52, 40-53, 40-54, 40-55, 40-56, 40-

[0166] 57, 40-58, 40-59, 40-60, 40-61 , 40-62, 40-63, 40-64, 40-65, 40-66, 40-67, 40-68, 40-69 or 40-

[0167] 70 nucleotides (or base pairs) in length.

[0168] Suitably, in some embodiments, the oligonucleotide may be between 45-46, 45-47, 45-48, 45- 49, 45-50, 45-51 , 45-52, 45-53, 45-54, 45-55, 45-56, 45-57, 45-58, 45-59, 45-60, 45-61 , 45- 62, 45-63, 45-64, 45-65, 45-66, 45-67, 45-68, 45-69, 45-70 nucleotides (or base pairs) in length.

[0169] In some embodiments, the oligonucleotide is between 35 to 70 base pairs (or nucleotides) in length. In some embodiments, the oligonucleotide is between 40 to 70 base pairs (or nucleotides) in length.

[0170] In some embodiments, the oligonucleotide is an shRNA and is 40 nucleotides in length. In some embodiments, the oligonucleotide is an shRNA and is 50 nucleotides in length. In some embodiments, the oligonucleotide is an shRNA and is 60 nucleotides in length. In some embodiments, the oligonucleotide is an shRNA and is 70 nucleotides in length.

[0171] Capability of binding & region of complementarity

[0172] The oligonucleotides described herein are capable of binding to a target sequence, suitably the oligonucleotides are capable of binding to the target sequence through being sufficiently complementary to the target sequence to allow hybridisation to occur under physiological conditions. Suitably therefore, the oligonucleotides described herein are capable of hybridising to the target sequence.

[0173] Suitably therefore, an oligonucleotide of the invention comprises or consists of a region of complementarity to the target sequence. The term “complementary” as used herein means a relationship in which nucleobases can form so-called Watson-Crick base pairs (natural type base pairs), or non-Watson-Crick base pairs (Hoogsteen type base pairs, or the like) via hydrogen bonds. Typically A is considered complementary to T, and G is considered complementary to C. Complementarity is expressed in terms of the number of residues that are able to form base pairs expressed as a proportion of some length. For example, if an oligonucleotide is 50% complementary to a target sequence this means 50% of the nucleotides comprised on one strand of that oligonucleotide (whether that oligonucleotide is single-stranded such as an ASO or shRNA or double-stranded such as an siRNA) are complementary to said target sequence.

[0174] In some embodiments, the entire length of the oligonucleotide may be complementary to the target sequence. In other words, the entire length of the oligonucleotide from it’s 5’ end to it’s 3’ end may constitute the region of complementarity, in other words the region of complementarity may comprise or consist of the entire length of the oligonucleotide from it’s 5’ end to it’s 3’ end. Suitably, antisense oligonucleotides (ASOs) and small interfering RNAs (siRNAs) are typically complementary across their entire length to a target sequence. Suitably, siRNAs are typically complementary across the entire length of their guide strand.

[0175] Alternatively, about half of the entire length of the oligonucleotide may be complementary to the target sequence. Suitably, short hairpin RNAs (shRNAs) are hairpin structures whereby typically about half the length of the oligonucleotide is complementary to a target sequence.

[0176] Suitably therefore, the region of complementarity may comprise part of the length of the oligonucleotide, suitably covering at least 40%, 41 %, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 60% 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, at least 99%, or up to 100% of the total length of the oligonucleotide. For example, a central portion of the oligonucleotide such as an ASO or siRNA may be complementary to the target sequence with the flanking terminal portions being non-complementary. Or for example, only about one half of the oligonucleotide such as an shRNA may be complementary to the target sequence with the other half being complementary to the first half wherein the first two halves are separated by a short loop region (allowing a hairpin structure to form) of less than 10, 9, 8, 7, 6, 5 or less than 4 nucleotides.

[0177] Similarly, it is understood that in order for the oligonucleotide to be capable of binding to the target sequence, it does not require that the entire length of the oligonucleotide to be capable of binding to the target sequence. It will be appreciated that a portion of the oligonucleotide may not be capable of binding to the target sequence, for example one or more nucleotides at the 5' or the 3' ends of an oligonucleotide such as an ASO or siRNA. Suitably therefore the oligonucleotide may comprise a region of complementarity and one or more nucleotides at the 5’ or 3’ end thereof which are non-complementary to the target sequence. Suitably therefore the oligonucleotide may comprise a region of complementarity and one or more flanking regions, suitably at the 5’ and / or 3’ end thereof, which are non-complementary to the target sequence.

[0178] Suitably, the region of complementarity may be at least 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20 or more nucleotides (or base pairs) in length. Suitably, the region of complementarity may be at least 5 or more nucleotides (or base pairs) in length. Suitably, the region of complementarity may be at least 10 or more nucleotides (or base pairs) in length. Suitably, the region of complementarity may be at least 15 or more nucleotides (or base pairs) in length. Suitably, the region of complementarity may be at least 20 or more nucleotides (or base pairs) in length.

[0179] Suitably, the region of complementarity may be up to 120, 110, 100, 90, 80, 70, 60, 50, 40, 30, 25, 23, 20 or less nucleotides (or base pairs) in length. Suitably, the region of complementarity may be up to 60 nucleotides (or base pairs) or less in length. Suitably, the region of complementarity may be up to 50 nucleotides (or base pairs) or less in length. Suitably, the region of complementarity may be up to 40 nucleotides (or base pairs) or less in length. Suitably, the region of complementarity may be up to 30 nucleotides (or base pairs) or less in length. Suitably, the region of complementarity may be up to 25 nucleotides (or base pairs) or less in length. Suitably, the region of complementarity may be up to 23 nucleotides (or base pairs) or less in length. Suitably, the region of complementarity may be up to 20 nucleotides (or base pairs) or less in length.

[0180] In one embodiment, the region of complementarity is 20 nucleotides (or base pairs) in length. In one embodiment, the oligonucleotide is 20 nucleotides (or base pairs) in length and comprises a region of complementarity to the target sequence that is 20 nucleotides (or base pairs) in length.

[0181] In one embodiment, the region of complementarity is 25 nucleotides (or base pairs) in length. In one embodiment, the oligonucleotide is 25 nucleotides (or base pairs) in length and comprises a region of complementarity to the target sequence that is 25 nucleotides (or base pairs) in length. In one embodiment, the oligonucleotide is an siRNA comprising a guide strand which is 25 nucleotides in length and comprises a region of complementarity to the target sequence that is 25 nucleotides in length. In one embodiment, the region of complementarity is anywhere from 18 to 23 nucleotides (or base pairs) in length. In one embodiment, the oligonucleotide is anywhere from 18 to 23 nucleotides (or base pairs) in length and comprises a region of complementarity to the target sequence that is the same length as the length of the nucleotide, suitably wherein the region of complementarity is anywhere from 18 to 23 nucleotides (or base pairs) in length.

[0182] The region of complementarity may be partially or fully complementary to the target sequence. Suitably therefore, the oligonucleotide comprises or consists of a region of complementarity to the target sequence, suitably wherein the complementarity may be partial or complete complementarity.

[0183] Complete complementarity refers to the entire region of complementary being 100% complementary to the target sequence (in other words, 100% of the reverse complement sequence of the region of complementarity being identical to the target sequence).

[0184] Partial complementarity refers to less than 100% complementarity, suitably the region of complementarity being at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or at least 99% complementary to the target sequence.

[0185] Suitably, the region of complementarity may comprise one or more, such as up to 60, 59, 58, 57, 56, 55, 54, 53, 52, 51 , 50, 49, 48, 47, 46, 45, 44, 43, 42, 41 , 40, 39, 38, 37, 36, 35, 34, 33, 32, 31 , 30, 29, 28, 27, 26, 25, 24, 23, 22, 21 , 20, 19, 18, 17, 16, 15, 14, 13, 12, 11 , 10, 9, 8, 7, 6, 5, 4, 3, 2 or up to 1 mismatched nucleotides when aligned or hybridised to the target sequence. For example, a 20 nt long oligonucleotide such as an ASO may have up to 10 nt that are mismatched when aligned or hybridised to the target sequence and this would represent 50% complementarity between the oligonucleotide and the target sequence.

[0186] The number of mismatches which may be tolerated without compromising the oligonucleotide’s ability of binding to the MEG3 transcript and reducing the level of MEG3 expression in a cell may be determined using routine in vitro assays measuring MEG3 expression when cells are treated with said oligonucleotide, such as those assays detailed in the Examples.

[0187] Single-Zdouble-stranded oligonucleotide

[0188] Suitably, the oligonucleotide may be single-stranded, double-stranded or comprise both single-stranded and double-stranded regions. For example, a single-stranded oligonucleotide may comprise double-stranded regions, and a double-stranded oligonucleotide may comprise single-stranded regions. In one embodiment, the oligonucleotide is single-stranded. In one embodiment, the oligonucleotide is double-stranded.

[0189] As is known in the art, an shRNA may be considered a single-stranded oligonucleotide which forms a hairpin structure to comprise a self-annealed double-stranded region with a singlestranded loop region.

[0190] In embodiments where the oligonucleotide is an shRNA or ASO (e.g. a mixmer ASO or gapmer ASO), the oligonucleotide is single-stranded. In embodiments where the oligonucleotide is a short interfering RNA (siRNA), the oligonucleotide is double-stranded.

[0191] In embodiments where the oligonucleotide is an ASO, the oligonucleotide is single-stranded and typically comprises or consist of deoxyribonucleotides, optionally further comprising ribonucleotides and / or modified nucleotides and / or non-naturally occurring derivatives, analogues or substitutes of nucleotides such as morpholino.

[0192] In embodiments where the oligonucleotide is an shRNA, the oligonucleotide is single-stranded and typically comprises or consist of ribonucleotides, optionally further comprising deoxyribonucleotides and / or modified nucleotides and / or non-naturally occurring derivatives, analogues or substitutes of nucleotides such as morpholino.

[0193] In embodiments where the oligonucleotide is an siRNA, the oligonucleotide is double-stranded (optionally comprising single-stranded regions, e.g., where the two strands of an siRNA have different lengths) and typically comprises or consist of ribonucleotides, optionally further comprising deoxyribonucleotides and / or modified nucleotides and / or non-naturally occurring derivatives, analogues or substitutes of nucleotides such as morpholino.

[0194] Nucleotides of the oligonucleotide

[0195] The oligonucleotide may comprise or consist of any type of nucleotides. Suitable types of nucleotides which may be comprised in the oligonucleotide are as follows: ribonucleotides (RNA), deoxyribonucleotides (DNA) and / or non-naturally occurring derivatives, analogues or substitutes of nucleotides such as morpholinos.

[0196] The oligonucleotide may comprise or consist of ribonucleotides (RNA). Suitably, the oligonucleotide may comprise or consist of deoxyribonucleotides (DNA). Suitably, the oligonucleotide may comprise or consist of both ribonucleotides (RNA) and deoxyribonucleotides (DNA). Suitably, the oligonucleotide may comprise or consist of non- naturally occurring derivatives, analogues or substitutes of nucleotides such as morpholines, suitably in combination with ribonucleotides and / or deoxyribonucleotides.

[0197] In some embodiments, the oligonucleotide comprises or consists of deoxyribonucleotides, optionally further comprising ribonucleotides and / or derivatives, analogues or substitutes of nucleotides such as morpholines, suitably wherein the oligonucleotide is an ASO. In one embodiment the oligonucleotide is an ASO comprising deoxyribonucleotides and ribonucleotides.

[0198] In some embodiments, the oligonucleotide comprises or consist of ribonucleotides, optionally further comprising deoxyribonucleotides and / or derivatives, suitably wherein the oligonucleotide is an siRNA or shRNA. In one embodiment, the oligonucleotide is an siRNA or shRNA consisting of ribonucleotides.

[0199] Modified nucleotides

[0200] Types of modified nucleotides

[0201] An oligonucleotide comprises nucleotides, which in turn typically consist of a nucleobase, a sugar (or derivative, analogue or substitute thereof) and an internucleoside linkage group.

[0202] Nucleotides of the oligonucleotide may comprise nucleobases including but not limited to purines and pyrimidines, more specifically guanine (G), adenine, (A), cytosine, (C), thymine, (T) and uracil (II), as well as natural or non-natural derivatives and modified or artificial analogues thereof.

[0203] Nucleotides of the oligonucleotide may comprise sugars including but not limited to the naturally occurring pentose deoxyribose (which forms DNA) and ribose (which forms RNA), as well as non-naturally occurring derivatives or analogues thereof such as morpholine rings.

[0204] Nucleotides of the oligonucleotide may comprise internucleoside linkages including but not limited to phosphate groups (i.e. phosphodiester linkages) as well as several other derivatives or analogues thereof such as phosphorothioate groups, boranophosphate groups, and more as described elsewhere herein. Such groups may also be attached on the 5’ or 3’ termini of the oligonucleotide.

[0205] Suitably the oligonucleotide may comprise one or more modified nucleotides. Modifications may be to the nucleobase, sugar (or equivalent) or internucleoside linkage of a nucleotide. Nucleotide modifications may be selected from any modifications known in the art. Suitably therefore, the oligonucleotide may comprise any one or more modifications known in the art.

[0206] Suitably, the oligonucleotide may comprise one or more 2’-modified nucleotides, 2-modified nucleotides, 3-modified nucleotides, 4-modified nucleotides, 5-modified nucleotides, 6- modified nucleotides, 7-modified nucleotides and / or 8-modified nucleotides.

[0207] Suitably, 2’-modified nucleotides may be selected from the group consisting of: 2’-O-methyl (2’OMe), 2’-O-methoxyethyl (2’MOE), 2’-fluoro (2’F), 2’-O-aminopropyl (2’OAP), 2’-O- dimethylaminoethyl (2’ODMAE), 2’-O-dimethylaminopropyl (2’ODMAP), 2’-O- dimethylaminoethyloxyethyl (2’ODMAEOE), 2’-O-N-methylacetamido (2’ONMA) 2'- dimethylaminooxyethoxy (2’-DMAO), 2’-dimethylaminoethoxyethoxy (2’-DMAEOE) and 2’ amino nucleotides.

[0208] Suitably, 2-modified nucleotides may be selected from the group consisting of: 2-amino (such as 2-amino pyrimidines such as 2-aminopyrimidine, 2-aminothymine, 2-aminocytosine and 2- aminouracil), 2-hydroxyl (such as 2-hydroxyl purines such as 2-hydroxypurine, 2- hydroxyadenine and 2-hydroxyguanine), 2-propyl, 2-thio, 2-fluoro (such as 2-fluoro pyrimidines such as 2-fluoropyrimidine, 2-fluorothymine, 2-fluorocytosine and 2-fluorouracil) nucleotides.

[0209] Suitably, 3-modified nucleotides may be 3-deaza nucleotides.

[0210] Suitably, 4-modified nucleotides may be 4-thio nucleotides such as 4-thio pyrimidines such as 4-thiopyrimidine, 4-thiothymine, 4-th iocytosine and 4-thiouracil.

[0211] Suitably, 5-modified nucleotides may be selected from the group consisting of: 5-methyl nucleotides (such as 5-methylcytosine, i.e. 5MeC), 5-halo, 5-propynyl, 5- hydroxymethyl, 5- bromo and 5-trifluoromethyl nucleotide nucleotides.

[0212] Suitably, 6-modified nucleotides may be 6-methyl or 6-azo nucleotides.

[0213] Suitably, 7-modified nucleotides may be 7-methyl or 7-deaza nucleotides.

[0214] Suitably, 8-modified nucleotides may be selected from the group consisting of: 8-halo, 8-amino, 8-thiol, 8-thioalkyl and 8-hydroxyl nucleotides.

[0215] Suitably, modified nucleobases may further include xanthine and hypoxanthine, and modified versions thereof. Suitably, the oligonucleotide may comprise one or more nucleotides with substituted sugar groups (i.e. where one or more groups of the sugar are substituted), suitably selected from the group comprising: OH; F; O-, S-, or N-alkyl; O-, S-, or N-alkenyl; O-, S- or N-alkynyl; or O-alkyl-O-alkyl, wherein the alkyl, alkenyl and alkynyl may be substituted or unsubstituted Ci to Cw alkyl or C2to Cw alkenyl and alkynyl. Suitably, these may include O[(CH2)nO]mCH3, O(CH2)nOCH3, O(CH2)nNH2, O(CH2)nCH3, O(CH2)nONH2, and O(CH2)nON[(CH2)nCH3]2, where n and m are from 1 to about 10.

[0216] Suitably, the oligonucleotide may comprise one or more substituted sugar groups selected from the group comprising: Ci to C lower alkyl, substituted lower alkyl, alkenyl, alkynyl, alkaryl, aralkyl, O-alkaryl or O-aralkyl, SH, SCH3, OCN, Cl, Br, CN, CF3, OCF3, SOCH3, SO2CH3, ONO2, NO2, N3, NH2, heterocycloalkyl, heterocycloalkaryl, aminoalkylamino, polyalkylamino, substituted silyl, methoxy, aminopropoxy, -O-allyl, an RNA cleaving group, a reporter group, an intercalator, a group for improving the pharmacokinetic properties of an oligonucleotide, or a group for improving the pharmacodynamic properties of an oligonucleotide, and other substituents having similar properties.

[0217] In one embodiment, the oligonucleotide may comprise one or more 2’-O-methoxyethyl (2’MOE) and one or more 5-methylcytosine (5MeC) nucleotides.

[0218] Suitably, the oligonucleotide may comprise one or more 2’-4’ bicyclic modified nucleotides, suitably selected from the group consisting of: locked nucleotides (locked nucleic acid monomers i.e. LNA monomers), 2’, 4’-constrained 2’-O-methoxyethyl (c2’MOE) nucleotides, 2’, 4’-constrained 2’-0-ethyl (cEt) nucleotides, and 2’-O, 4’-C-ethylene-bridged (ENA) nucleotides (a.k.a. bridged nucleic acids), tricyclo nucleotides.

[0219] Suitably, the oligonucleotide may comprise one or more artificial nucleotides suitably selected from the group consisting of: peptide nucleic acids (PNAs, i.e. peptide nucleotides) and phosphorodiamidate morpholino monomers. In a PNA monomer, the sugar-phosphate backbone of a nucleotide is substituted with a pseudo-peptide polymer to which the nucleobases are linked. In a phosphorodiamidate morpholino monomer, the sugar groups of a nucleotide are replaced with a morpholine ring and the internucleoside linkages of a nucleotide are a phosphorodiamidate group.

[0220] Suitably the oligonucleotide may consist of PNAs, suitably the oligonucleotide may be a peptide nucleic acid, e.g. a peptide oligonucleic acid. Suitably the oligonucleotide may consist of phosphorodiamidate morpholino monomers, suitably the oligonucleotide may be a phosphorodiamidate morpholino oligomer (PMO).

[0221] Suitably, the oligonucleotide may comprise one or more modified internucleoside linkages (i.e. internucleoside linkage groups), suitably selected from the group consisting of: a phosphotriester linkage, a phosphorothioate (PS) internucleoside linkage, a methylphosphonate internucleoside linkage, a phosphorodithioate internucleoside linkage, a boranophosphate internucleoside linkage, a phosphoramidate internucleoside linkage, a phosphorodiamidate internucleoside linkage, a thiophosphoramidate linkage, a thiophosphorodiamidate linkage, a phosphonocarboxylate internucleoside linkage, a phosphonoacetate internucleoside linkage, an aminoalkylphosphotriester internucleoside linkage, a methyl or other alkyl phosphonate internucleoside linkage (such as a 3’-alkylene phosphonate internucleoside linkage or a 5’-alkylene phosphonate internucleoside linkage), a phosphinate internucleoside linkage, a phosphoramidate internucleoside linkage (such as a 3’-amino phosphoramidate internucleoside linkage or an aminoalkylphosphoramidate internucleoside linkage), a thionophosphoramidate internucleoside linkage, a thionoalkylphosphotriester internucleoside linkage, a selenophosphate internucleoside linkage.

[0222] Suitably, one or more modified internucleoside linkages may be 3’-5’ internucleoside linkages, 2’-5’ internucleoside linkages, 2’-2’ internucleoside linkages, 3’-3’ internucleoside linkages or 5’-5’ internucleoside linkages. Suitably, one or more modified internucleoside linkages may be inverted polarity internucleoside linkages.

[0223] In one embodiment, the oligonucleotide may comprise one or more phosphorothioate (PS) internucleoside linkages.

[0224] Suitably the oligonucleotide may comprise one or more asymmetric centres and thus give rise to enantiomers, diasteromers, and other stereoisomeric configurations, e.g. R, S. For example, stereochemistry at the phosphorus atom in the internucleoside linkage may be manipulated to produce phosphorothioate (PS) linkages with controlled R and S configurations. Stereochemistry may be constrained at one or more modified internucleoside linkages. This stereochemical precision can reduce off-target effects and improve the pharmacokinetic and pharmacodynamic profiles of the oligonucleotides. Suitably the oligonucleotide is at least 10%, 20%, 30%, 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99% diastereomerically pure. The oligonucleotide may comprise one or more chiral internucleoside linkages. The use of chiral internucleosidic linkages in oligonucleotide synthesis can create a chiral centre at the phosphorus linkage. A chiral linkage phosphorus centre can have either an “Sp” or “Rp” configuration. A conventional stereorandom preparation of an oligonucleotide that contains n chiral linkage phosphorus is a mixture of 2n stereoisomers with respect to chiral linkage phosphorus centres, each of which stereoisomers share the same constitution but differs in stereochemistry along its backbone. Without control of stereochemistry of backbone chiral centres, a stereorandom oligonucleotide preparation (e.g., a random mixture of diastereoisomers) provides uncontrolled (or stereorandom) compositions comprising undetermined levels of oligonucleotide stereoisomers. Even though these stereoisomers may have the same base sequence and / or chemical modifications, they are different chemical entities at least due to their different backbone stereochemistry, and they can have different properties, e.g., activities, toxicities, distribution etc. In some embodiments, each chiral internucleosidic linkage is independently formed with about 97% or more diastereoselectivity (e g., as measured through preparation of a suitable dimer). In some embodiments, most chiral internucleosidic linkages are independently formed with about 98% or more diastereoselectivity. In some embodiments, one or more, e.g., 1 , 2, 3, 4, 5, 6, 7, 8 or more chiral internucleosidic linkages are independently formed with about 99% or more diastereoselectivity. In some embodiments, overall diastereoselectivity (as product of diastereoselectivity of all chiral internucleosidic linkages) is about 80% or more. In some embodiments, it is about 81 % or more. In some embodiments, it is about 82% or more. In some embodiments, it is about 83% or more. In some embodiments, it is about 84% or more. In some embodiments, it is about 85% or more. In some embodiments, the oligonucleotide is stereodefined or stereopure. In some embodiments, the oligonucleotide may therefore be referred to as chirally controlled. In some embodiments, level of a particular stereoisomer, of a chirally controlled oligonucleotide composition is enriched as described herein (e.g., in some embodiments, each chiral internucleosidic linkage independently has a stereopurity of about 97%, 98%, 99% or more). In some embodiments, each chirally controlled internucleoside linkage independently has a diastereopurity of at least 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% with respect to its chiral linkage phosphorus.

[0225] Suitably, modified nucleotides may comprise any other modification known in the art.

[0226] Combinations of modified nucleotides

[0227] The oligonucleotide may comprise a single type of nucleotide modification described herein or combinations of different nucleotide modifications described herein. Suitably any given nucleotide may comprise multiple different modifications as described herein. For example, nucleotides may be 2’-O-methoxyethyl (2’MOE) modified 5-methylcytosines (5MeC).

[0228] In some embodiments, the oligonucleotide comprises one or more 2’-O-methoxyethyl (2’MOE) nucleotides. In some embodiments, the oligonucleotide comprises one or more 5- methylcytosine (5MeC) nucleotides. In some embodiments, the oligonucleotide comprises one or more 2’-O-methoxyethyl (2’MOE) nucleotides and one or more 5-methylcytosine (5MeC) nucleotides. In some embodiments, the oligonucleotide comprises one or more 2’-O- methoxyethyl (2’MOE) modified 5-methylcytosines (5MeC), suitably in addition to one or more 2’-O-methoxyethyl (2’MOE) nucleotides and / or one or more 5-methylcytosine (5MeC) nucleotides. In one embodiment, the oligonucleotide comprises one or more 2’-O- methoxyethyl (2’MOE) nucleotides, one or more 5-methylcytosine (5MeC) nucleotides and one or more 2’-O-methoxyethyl (2’MOE) modified 5-methylcytosines (5MeC).

[0229] In some embodiments, the oligonucleotide comprises one or more phosphorothioate internucleoside linkages. In some embodiments, each internucleoside linkage of the oligonucleotide is a phosphorothioate linkage (i.e. a fully PS oligonucleotide).

[0230] In some embodiments, the oligonucleotide comprises one or more phosphorothioate internucleoside linkages, and any one or more of: one or more 2’-O-methoxyethyl (2’MOE) nucleotides, one or more 5-methylcytosine (5MeC) nucleotides, and one or more 2’-O- methoxyethyl (2’MOE) modified 5-methylcytosines (5MeC).

[0231] In some embodiments, each internucleoside linkage of the oligonucleotide is a phosphorothioate linkage, and the oligonucleotide further comprises one or more 2’-O- methoxyethyl (2’MOE) nucleotides, one or more 5-methylcytosine (5MeC) nucleotides, and / or one or more 2’-O-methoxyethyl (2M’OE) modified 5-methylcytosines (5MeC).

[0232] In one embodiment, each internucleoside linkage of the oligonucleotide is a phosphorothioate linkage, and the oligonucleotide further comprises one or more 2’-O-methoxyethyl (2’MOE) nucleotides, one or more 5-methylcytosine (5MeC) nucleotides, and one or more 2’-O- methoxyethyl (2’MOE) modified 5-methylcytosines (5MeC).

[0233] In some embodiments, the oligonucleotide comprises deoxyribonucleotides comprising one or more 5-methylcytosines (5MeC). In some embodiments, the oligonucleotide comprises deoxyribonucleotides wherein each cytosine deoxyribonucleotide is a 5-methylcytosine (5MeC) (i.e. wherein each cytosine deoxyribonucleotide is 5-methyl modified). In some embodiments, the oligonucleotide comprises ribonucleotides comprising one or more 2’-O-methoxyethyl (2’MOE) modified nucleotides. In some embodiments, the oligonucleotide comprises ribonucleotides consisting of 2’-O-methoxyethyl (2’MOE) modified nucleotides. In some embodiments, the oligonucleotide comprises ribonucleotides consisting of 2’-O- methoxyethyl (2’MOE) modified nucleotides, wherein each cytosine ribonucleotide is also 5- methylcytosine (5MeC) (i.e. wherein each cytosine ribonucleotide is also 5-methyl modified, i.e. is a 2’MOE 5MeC).

[0234] In one embodiment, the oligonucleotide comprises deoxyribonucleotides wherein each cytosine deoxyribonucleotide is a 5-methylcytosine (5MeC), and ribonucleotides consisting of 2’-O-methoxyethyl (2’MOE) modified nucleotides wherein each cytosine ribonucleotide is also a 5-methylcytosine (5MeC) (i.e. wherein each cytosine in the oligonucleotide whether ribonucleotide or deoxyribonucleotide is 5-methyl modified), and wherein the oligonucleotide comprises one or more phosphorothioate internucleoside linkages.

[0235] In one embodiment, the oligonucleotide comprises deoxyribonucleotides wherein each cytosine deoxyribonucleotide is a 5-methylcytosine (5MeC), and ribonucleotides consisting of 2’-O-methoxyethyl (2’MOE) modified nucleotides wherein each cytosine ribonucleotide is also a 5-methylcytosine (5MeC) (i.e. wherein each cytosine in the oligonucleotide whether ribonucleotide or deoxyribonucleotide is 5-methyl modified), and wherein each internucleoside linkage of the oligonucleotide is a phosphorothioate linkage (i.e. the oligonucleotide is a fully phosphorothioated oligonucleotide).

[0236] Nucleotide modifications such as 2'-MOE modifications are known to amongst other things enable enhanced binding affinity to a target nucleic acid with minimal toxicity. Modified internucleoside linkage groups such as PO and PS modifications are known to result in resistance to a broad spectrum of nucleases and increase protein binding, which also improves tissue uptake, and may be useful for fine tuning the pharmacokinetics of the oligonucleotide. A 5-methylcytosine substitution reduces ASO immunostimulatory effects.

[0237] Types of oligonucleotides

[0238] The oligonucleotide may any type of oligonucleotide known in the art such as an antisense oligonucleotides (ASO), a short interfering RNAs (siRNA), a short hairpin RNAs (shRNA), an anti-miRNAs (antimiR), a splice-switching oligonucleotide, a DNA or RNA aptamer, and more.

[0239] In some embodiments, the oligonucleotide is an ASO.

[0240] In some embodiments, the oligonucleotide is an siRNA. In some embodiments, the oligonucleotide is an shRNA.

[0241] In one preferred embodiment, the oligonucleotide is an ASO, suitably a gapmer ASO as defined hereinbelow.

[0242] The present invention includes any pharmaceutically acceptable salt of the oligonucleotide, an ester of the oligonucleotide, or a salt of the ester. Suitably, the present invention includes any pharmaceutically acceptable salt of, an ester of, or a salt of the ester of the oligonucleotide (such as the gapmer or mixmer ASO, siRNA or shRNA) of the invention. A person skilled in the art will be aware of pharmaceutically acceptable salts of oligonucleotides. Examples of suitable pharmaceutically acceptable salts include, but are not limited to, a sodium salt, a potassium salt, and a meglumine salt.

[0243] Regardless of the type of the oligonucleotide, the oligonucleotide may comprise or consist of a sequence with at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity to a sequence selected from the group consisting: SEQ ID NO: 10-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 10, 14-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 10, 14, 17-40.

[0244] Suitably, the oligonucleotide may comprise or consist of a sequence with at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity to a sequence selected from the group consisting of: SEQ ID NO: 17-40, suitably to a sequence selected from the group consisting of: SEQ I D NO: 17, 20-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 17, 20-27, 29-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 17, 21-27, 29-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 20-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 21-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 20-27, 29-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 21-27, 29-40.

[0245] Suitably, the oligonucleotide may comprise or consist of a sequence with at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity to a sequence selected from the group consisting of: SEQ ID NO: 23-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 23-27, 29-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 23-25, 27, 29-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 23-27, 29-37, 39-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 23-25, 27, 29-37, 39-40, suitably to a sequence selected from the group consisting of: SEQ ID NO: 23-25, 27, 29-33, 35-37, 39-40, suitably to a sequence selected from the group consisting of: 23-25, 27, 29-32, 35-37, 39-40.

[0246] The oligonucleotide may comprise a functional variant of any of the sequences mentioned above, suitably any of SEQ ID NO: 10-40. By “functional variant” is meant a variant which maintains it’s functionality in being able to reduce the level of MEG3 expression, suitably in by binding to MEG3. Functional variants may be determined by a skilled person using techniques described in the Examples. Functional variants may include modifications such as truncations or a small number of insertions, deletions and / or substitutions in a given sequence, suitably between 1-10 nucleotide modifications. Merely by way of an example, a functional variant of SEQ ID NO: 10 may include a single nucleotide substitution or for example a trinucleotide deletion in the sequence of SEQ ID NO: 10.

[0247] Antisense

[0248] The oligonucleotide may be an ASO, suitably wherein the ASO may be a mixmer ASO or a gapmer ASO. A gapmer ASO comprises a central region or ‘gap’ flanked on either side by a first and a second flanking region. In other words, a gapmer ASO comprises the structure 5’- A-B-C-3’, wherein B is the central region and A and C are the flanking regions. In one embodiment, the oligonucleotide is a gapmer ASO.

[0249] Suitably the central region may be complementary to the target sequence, suitably the central region may comprise the region of complementarity. In addition, the one or both flanking regions may also be complementary to the target sequence, suitably one or both flanking regions may also comprise the region of complementarity. Suitably the region of complementarity may comprise the central (B) and both flanking regions (A and C). Suitably the region of complementarity may extend across the entire sequence of the ASO.

[0250] Suitably, the structure of an oligonucleotide may comprise of an asymmetrical format (i.e. design). Suitably, the structure of an oligonucleotide may comprise of a symmetrical format. Suitably therefore, in some embodiments the gapmer ASO may comprise flanking regions of different lengths, and comprising different modifications. Suitably therefore, in some embodiments the gapmer ASO may comprise flanking regions of the same lengths, and comprising the same modifications in corresponding positions. In one embodiment, the ASO is a gapmer comprising a symmetrical format.

[0251] Suitably, the central region of a gapmer ASO may comprise or consist of deoxyribonucleotides or ribonucleotides, suitably deoxyribonucleotides, which may be modified or unmodified. Suitably, the flanking regions of a gapmer ASO may each independently comprise or consist of deoxyribonucleotides or ribonucleotides, suitably ribonucleotides, which may be modified or unmodified. In some embodiments, the oligonucleotide is a gapmer ASO comprising a central region consisting of deoxyribonucleotides and flanking regions consisting of ribonucleotides.

[0252] Suitably therefore, a gapmer ASO may comprise the structure 5’-A-B-C-3’, wherein A, B and C each independently comprise or consist of ribonucleotides or deoxyribonucleotides, suitably wherein A and C consist of ribonucleotides or deoxyribonucleotides, and B consists of deoxyribonucleotides. Suitably wherein A and C consist of ribonucleotides, and B consists of deoxyribonucleotides.

[0253] Suitably the central region (B) may comprise up to 80 nucleotides, and the flanking regions (A and C) may comprise up to 20 nucleotides. In some embodiments, the oligonucleotide is a gapmer ASO comprising a central region (B) consisting of between 5-30, 5-20 or 7-12 (all ranges mentioned herein include the terminal values) deoxyribonucleotides and flanking regions (A and C) each independently consisting of between 1-10, 3-7 or 3-5 ribonucleotides. In some embodiments, the oligonucleotide is a gapmer ASO comprising a central region (B) consisting of 10 (or about 10) deoxyribonucleotides and flanking regions (A and C) each consisting of 5 (or about 5) ribonucleotides.

[0254] Nucleotides in the central and flanking regions may each independently comprise any number of any combination of any modification mentioned elsewhere herein. For example, the flanking regions and central regions may each independently comprise one or more 2’-O-methyl (2OMe) modified nucleotides, 2'-O-methoxyethyl (2 -MOE) modified nucleotides, locked nucleic acids (LNAs), 2, 4-constrained 2-0-ethyl (cEt) nucleotides and / or phosphorothioate (PS) internucleoside linkages.

[0255] In some embodiments, the central region consists of deoxyribonucleotides comprising one or more 5-methylcytosine (5MeC) nucleotides. In some embodiments, the central region consists of deoxyribonucleotides wherein each cytosine is a 5-methylcytosine (5MeC) (i.e. wherein each cytosine is 5-methyl modified). In some embodiments, the flanking regions consist of ribonucleotides comprising one or more 2’-O-methoxyethyl (2’MOE) modified nucleotides. In some embodiments, the flanking regions consist of ribonucleotides consisting of 2’-O-methoxyethyl (2’MOE) modified nucleotides. In some embodiments, the flanking regions consist of ribonucleotides consisting of 2’-O- methoxyethyl (2’MOE) modified nucleotides wherein each cytosine is also a 5-methylcytosine (5MeC) (i.e. is also 5-methyl modified).

[0256] In some embodiments, the oligonucleotide is a gapmer ASO comprising a central region (B) consisting of between 5-30, 5-20 or 7-12 deoxyribonucleotides comprising one or more 5- methylcytosine (5MeC) nucleotides, and flanking regions (A and C) consisting of between 1- 10, 3-7 or 3-5 ribonucleotides comprising one or more 2’-O-methoxyethyl (2’MOE) modified nucleotides wherein each cytosine is a 5-methylcytosine (5MeC).

[0257] In some embodiments, the oligonucleotide is a gapmer ASO comprising a central region (B) consisting of between 5-30, 5-20 or 7-12 deoxyribonucleotides wherein each cytosine is a 5- methylcytosine (5MeC), and flanking regions (A and C) consisting of between 1-10, 3-7 or 3- 5 ribonucleotides consisting of 2’-O-methoxyethyl (2’MOE) modified nucleotides wherein each cytosine is also a 5-methylcytosine (5MeC) (i.e. wherein each cytosine in the oligonucleotide whether ribonucleotide or deoxyribonucleotide is 5-methyl modified).

[0258] In one embodiment, the oligonucleotide is a gapmer ASO comprising a central region (B) consisting of 10 (or about 10) deoxyribonucleotides wherein each cytosine is a 5- methylcytosine (5MeC), and flanking regions (A and C) each consisting of 5 (or about 5) ribonucleotides consisting of 2’-O-methoxyethyl (2’MOE) modified nucleotides wherein each cytosine is also a 5-methylcytosine (5MeC) (i.e. wherein each cytosine in the oligonucleotide whether ribonucleotide or deoxyribonucleotide is 5-methyl modified).

[0259] In one embodiment, the oligonucleotide is a gapmer ASO comprising a central region (B) consisting of 10 (or about 10) deoxyribonucleotides wherein each cytosine is a 5- methylcytosine (5MeC), and flanking regions (A and C) each consisting of 5 (or about 5) ribonucleotides consisting of 2’-O-methoxyethyl (2’MOE) modified nucleotides wherein each cytosine is also a 5-methylcytosine (5MeC) (i.e. wherein each cytosine in the oligonucleotide whether ribonucleotide or deoxyribonucleotide is 5-methyl modified), and wherein the oligonucleotide comprises one or more phosphorothioate internucleoside linkages.

[0260] In one embodiment, the oligonucleotide is a gapmer ASO comprising a central region (B) consisting of 10 (or about 10) deoxyribonucleotides wherein each cytosine is a 5- methylcytosine (5MeC), and flanking regions (A and C) each consisting of 5 (or about 5) ribonucleotides consisting of 2’-O-methoxyethyl (2’MOE) modified nucleotides wherein each cytosine is also a 5-methylcytosine (5MeC) (i.e. wherein each cytosine in the oligonucleotide whether ribonucleotide or deoxyribonucleotide is 5-methyl modified), and each internucleoside linkage of the oligonucleotide is a phosphorothioate linkage (i.e. the oligonucleotide is a fully phosphorothioated oligonucleotide). An advantage of such an ASO is that it does not exert any observable cytotoxicity as demonstrated in the Examples.

[0261] Optionally the oligonucleotide may comprise one or more terminal 5’ or 3’ modifications. Optionally the oligonucleotide may comprise a phosphate, inverted dT, cholesterol or other sterol, polyethylene glycol (PEG) or other polymer, a fluorescent label such as FAM or Cy5, biotin or other affinity tag, a methyl group, a fluorine, or an antibody, cell penetrating peptide, or GalNAc group at the 3’ or the 5’ end thereof, for example.

[0262] In one embodiment, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from any of (i.e. selected from the group consisting of) SEQ ID NO: 10-40, suitably a sequence selected from any of: SEQ ID NO: 10, 14-40, suitably a sequence selected from any of: SEQ ID NO: 10, 14, 17-40, suitably a sequence selected from any of: SEQ ID NO: 17-40, suitably a sequence selected from any of: SEQ ID NO: 17, 20-40, suitably a sequence selected from any of: SEQ ID NO: 17, 20-27, 29-40, suitably a sequence selected from any of: SEQ ID NO: 17, 21-27, 29-40, suitably a sequence selected from any of: SEQ ID NO: 20-40, suitably a sequence selected from any of: SEQ ID NO: 21- 40, suitably a sequence selected from any of: SEQ ID NO: 20-27, 29-40, suitably a sequence selected from any of: SEQ ID NO: 21-27, 29-40, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 57-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 57, 61-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 57, 61 , 64-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 64-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 64, 67-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 64, 67-74,76-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 64, 68-74,76-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 67-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 68-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 67-74, 76-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 68-74, 76-87, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0263] Suitably, the oligonucleotide may target SEQ ID NO: 1 (ENSE00002476600). Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from any one of SEQ ID NO: 10-16, suitably any one of SEQ ID NO: 10 and 14, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 57-63, suitably a sequence selected from the group consisting of: SEQ ID NO: 57 and 61 , or a sequence having at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0264] Suitably, the oligonucleotide may target SEQ ID NO: 2 (ENSE00001586203). Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting a sequence selected from any one of SEQ ID NO: 17-22, suitably any one of SEQ ID NO: 17, 20-22, suitably any one of SEQ ID NO: 17, 21 , 22, suitably any one of SEQ ID NO: 20-22, suitably any one of SEQ ID NO: 21 and / or 22, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 64-69, suitably a sequence selected from the group consisting of: SEQ ID NO: 64, 67-69, suitably a sequence selected from the group consisting of: SEQ ID NO: 64, 68, 69, suitably a sequence selected from the group consisting of: SEQ ID NO: 67-69, suitably a sequence selected from the group consisting of: SEQ ID NO: 68 and / or 69, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0265] Suitably, the oligonucleotide may target SEQ ID NO: 4. Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting a sequence selected from any one of SEQ ID NO: 21 and 22, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 68 and 69, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0266] Suitably, the oligonucleotide may target SEQ ID NO: 3 (ENSE00002440436). Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from any one of SEQ ID NO: 23-40, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that target any one or more of SEQ ID NO: 5-9 as mentioned hereinbelow (since SEQ ID NO: 5-9 are comprised within SEQ ID NO: 3). Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 70-87, and / or any sequences that target any one or more of SEQ I D NO: 5-9 as mentioned hereinbelow, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0267] Suitably, the oligonucleotide may target SEQ ID NO: 5. Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from any one of SEQ ID NO: 23-27, 29-40, suitably any one of SEQ ID NO: 23-25, 27, 29-40, suitably any one of SEQ ID NO: 23-27, 29-37, 39-40, suitably any one of SEQ ID NO: 23-25, 27, 29- 37, 39-40, suitably any one of SEQ ID NO: 23-25, 27, 29-33, 35-37, 39-40, suitably any one of SEQ ID NO: 23-25, 27, 29-32, 35-37, 39-40, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that target any one or more of SEQ ID NO: 6-9 as mentioned hereinbelow (since SEQ ID NO: 6-9 are comprised within SEQ ID NO: 5). Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 70-74, 76-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 70- 72, 74, 76-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 70-74, 76-84, 86-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 70-72, 74, 76-84, 86-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 70- 72, 74, 76-80, 82-84, 86-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 70-72, 74, 76-79, 82-84, 86-87, and / or any of those sequences that target any one or more of SEQ ID NO: 6-9 as mentioned hereinbelow, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0268] Suitably, the oligonucleotide may target SEQ ID NO: 6. Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from any one of SEQ I D NO: 23-25, 29-40, suitably any one of SEQ I D NO: 23-25, 29-37, 39-40, suitably any one of SEQ ID NO: 23-25, 29-33, 35-37, 39-40, suitably any one of SEQ ID NO: 23-25, 29-32, 35-37, 39-40, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that target any one or more of SEQ ID NO: 7-9 as mentioned hereinbelow (since SEQ ID NO: 7-9 are comprised within SEQ ID NO: 6), suitably in combination with any one or more of SEQ ID NO: 33, 34, 38. Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 70-72, 76-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 70-72, 76-84, 86-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 70-72, 76-80, 82-84, 86-87, suitably a sequence selected from the group consisting of: SEQ ID NO: 70-72, 76-79, 82-84, 86-87, and / or any of those sequences that target any one or more of SEQ ID NO: 7-9 as mentioned hereinbelow, suitably in combination with any one or more of SEQ ID NO: 80, 81 , 85, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0269] Suitably, the oligonucleotide may target SEQ ID NO: 7. Suitably may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from any one of SEQ ID NO: 23, 24, 29-32, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 70, 71 , 76-79, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0270] Suitably, the oligonucleotide may target SEQ ID NO: 8. Suitably may be an ASO, suitably gapmer ASO, comprising or consisting of a sequence selected from any one of SEQ ID NO: 35-37, or a sequence having at least 50%, at least 60%, at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 82-84, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0271] Suitably, the oligonucleotide may target SEQ ID NO: 9. Suitably, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from any one of SEQ ID NO: 25, 39, 40, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of: SEQ ID NO: 72, 86, 87, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0272] Suitably the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of SEQ ID NO: 27 or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably therefore, the oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of a sequence selected from the group consisting of SEQ ID NO: 74, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. The oligonucleotide may equally target any one or more of SEQ ID NO: 119, 120, 48-56, 121- 126.

[0273] Suitably, an oligonucleotide targeting SEQ ID NO: 119 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 7 and / or 8. Suitably, an oligonucleotide targeting SEQ ID NO: 120 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 8 and / or 9.

[0274] Suitably, an oligonucleotide targeting SEQ ID NO: 48 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1-3.

[0275] Suitably, an oligonucleotide targeting SEQ ID NO: 49 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 3.

[0276] Suitably, an oligonucleotide targeting SEQ ID NO: 50 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 4 and / or 3.

[0277] Suitably, an oligonucleotide targeting SEQ ID NO: 51 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 5.

[0278] Suitably, an oligonucleotide targeting SEQ ID NO: 52 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 4 and / or 5.

[0279] Suitably, an oligonucleotide targeting SEQ ID NO: 53 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 6.

[0280] Suitably, an oligonucleotide targeting SEQ ID NO: 54 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 4 and / or 6. Suitably, an oligonucleotide targeting SEQ ID NO: 55 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 7.

[0281] Suitably, an oligonucleotide targeting SEQ ID NO: 56 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 4 and / or 7.

[0282] Suitably, an oligonucleotide targeting SEQ ID NO: 51 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 5.

[0283] Suitably, an oligonucleotide targeting SEQ ID NO: 121 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1 , 2 and / or 5.

[0284] Suitably, an oligonucleotide targeting SEQ ID NO: 122 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1 , 2 and / or 6.

[0285] Suitably, an oligonucleotide targeting SEQ ID NO: 123 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1 , 2 and / or 7.

[0286] Suitably, an oligonucleotide targeting SEQ ID NO: 124 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1 , 2 and / or 119.

[0287] Suitably, an oligonucleotide targeting SEQ ID NO: 125 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 119.

[0288] Suitably, an oligonucleotide targeting SEQ ID NO: 126 may be an ASO, suitably a gapmer ASO, comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 120.

[0289] In one preferred embodiment, oligonucleotide may be an ASO, suitably a gapmer ASO, comprising or consisting of SEQ ID NO: 23-40, suitably comprising or consisting of SEQ ID NO: 70-87. The oligonucleotide may comprise a functional variant of any of the sequences mentioned above, suitably any of SEQ ID NO: 57-87. By “functional variant” is meant a variant which maintains it’s functionality in being able to reduce the level of MEG3 expression, suitably in by binding to MEG3. Functional variants may be determined by a skilled person using techniques described in the Examples. Functional variants may include modifications such as truncations or a small number of insertions, deletions and / or substitutions in a given sequence, suitably between 1-10 nucleotide modifications. Merely by way of an example, a functional variant of SEQ ID NO: 57 may include a single nucleotide substitution or for example a trinucleotide deletion in the sequence of SEQ ID NO: 57.

[0290] Short interferinq RNAs

[0291] The oligonucleotide may be an siRNA.

[0292] As mentioned hereinabove, in embodiments where the oligonucleotide is an siRNA, the oligonucleotide is double-stranded (optionally comprising single-stranded regions) and typically comprises or consist of ribonucleotides, optionally further comprising deoxyribonucleotides and / or modified nucleotides and / or non-naturally occurring derivatives, analogues or substitutes of nucleotides such as morpholino.

[0293] Suitably, the siRNA may be partially single-stranded. Suitably, the termini (i.e. ends) of the siRNA molecule may be single-stranded, i.e. the siRNA may comprise overhangs, suitably 5’ and / or 3’ overhangs. Suitably, the siRNA may comprise a 5’ overhang at one terminus of the siRNA molecule, suitably the siRNA may comprise 5’ overhangs at both termini of the siRNA molecule. Suitably, the siRNA may comprise a 3’ overhang at one terminus of the siRNA molecule, suitably the siRNA may comprise 3’ overhangs at both termini of the siRNA molecule. For example, the siRNA may comprise both a 5’ overhang and a 3’ overhang. In one embodiment, the siRNA comprises 3’ overhangs on both termini of the siRNA molecule.

[0294] Suitably, in embodiments where the siRNA is partially single-stranded, the single-stranded regions may comprise or consist of deoxyribonucleotides or ribonucleotides, suitably the single-stranded regions may consist of deoxynucleotides. Therefore, in some embodiments, the siRNA comprises 5’ and / or 3’ overhangs consisting of deoxynucleotides.

[0295] Suitably, the overhangs may be 1 , 2, 3, 4 or 5 nucleotides or longer in length. In one embodiment, the overhangs are 2 nucleotides in length. In one embodiment, the overhangs comprise or consist of two thymine nucleotides (TT), suitably two thymine deoxyribonucleotides. In one embodiment, the siRNA comprises 3’ overhangs on both termini of the siRNA molecule, wherein the overhangs comprise or consist of two thymine deoxyribonucleotides (TT). In one embodiment, the oligonucleotide is an siRNA comprising or consisting of SEQ ID NO: 127 and SEQ ID NO: 128. In such an embodiment, suitably the siRNA targets a region of the MEG3 transcript which comprises or consists of SEQ ID NO: 3 and / or 5, and / or which comprises or consists of the target site SEQ ID NO: 105. Suitably the siRNA forms a double-stranded oligonucleotide with 3’ TT overhangs on both termini of the molecule.

[0296] Suitably, the siRNA may be complementary to the target sequence. Suitably, the siRNA may comprise the region of complementarity. Suitably, the entire length of the siRNA may be complementary to the target sequence. Suitably, the entire length of the shortest or longest strand of the siRNA may be complementary to the target sequence. Suitably, only part of the length of the siRNA may be complementary to the target sequence.

[0297] Suitably, the structure of an oligonucleotide may comprise of an asymmetrical format (i.e. design). Suitably, the structure of an oligonucleotide may comprise of a symmetrical format. Suitably therefore, the siRNA may comprise two strands of different lengths, or two strands of the same length.

[0298] Suitably, the siRNA may comprise any number of any combination of any nucleotide modification mentioned elsewhere herein. For example, the siRNA may comprise one or more 2’-O-methyl (2OMe) modified nucleotides, 2'-O-methoxyethyl (2'-MOE) modified nucleotides, locked nucleic acids (LNAs), 2, 4-constrained 2-O-ethyl (cEt) nucleotides and / or phosphorothioate (PS) internucleoside linkages.

[0299] In some embodiments, the siRNA comprises one or more 5-methylcytosine (5MeC) nucleotides. In some embodiments, the siRNA comprises one or more 2’-O-methoxyethyl (2’MOE) modified nucleotides. In some embodiments, the siRNA comprises one or more LNAs. In some embodiments, the siRNA comprises one or more PS linkages. In some embodiments, all internucleoside linkages comprised in the siRNA are PS linkages (i.e. both strands of the siRNA may be fully phosphorothioated).

[0300] Optionally the oligonucleotide may comprise one or more terminal 5’ or 3’ modifications. Optionally the oligonucleotide may comprise a phosphate, inverted dT, cholesterol or other sterol, polyethylene glycol (PEG) or other polymer, a fluorescent label such as FAM or Cy5, biotin or other affinity tag, a methyl group, a fluorine, or an antibody, cell penetrating peptide, or GalNAc group at the 3’ or the 5’ end thereof, for example. In one embodiment, the oligonucleotide may be an siRNA, comprising or consisting of a sequence selected from any of (i.e. selected from the group consisting of) SEQ ID NO: 129- 159, suitably a sequence selected from any of: SEQ ID NO: 129, 133-159, suitably a sequence selected from any of: SEQ ID NO: 129, 133, 136-159, suitably a sequence selected from any of: SEQ ID NO: 136-159, suitably a sequence selected from any of: SEQ ID NO: 136, 139- 159, suitably a sequence selected from any of: SEQ ID NO: 136, 139-146, 148-159, suitably a sequence selected from any of: SEQ ID NO: 136, 140-146, 148-159, suitably a sequence selected from any of: SEQ ID NO: 139-159, suitably a sequence selected from any of: SEQ ID NO: 140-159, suitably a sequence selected from any of: SEQ ID NO: 139-146, 148-159, suitably a sequence selected from any of: SEQ ID NO: 140-146, 148-159, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, suitably at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0301] Suitably, the oligonucleotide may target SEQ ID NO: 1 (ENSE00002476600). Suitably, the oligonucleotide may be an siRNA comprising or consisting of a sequence selected from any one of SEQ ID NO: 129-135, suitably any one of SEQ ID NO: 129 and 133, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0302] Suitably, the oligonucleotide may target SEQ ID NO: 2 (ENSE00001586203). Suitably, the oligonucleotide may be an siRNA comprising or consisting a sequence selected from any one of SEQ ID NO: 136-141 , suitably any one of SEQ ID NO: 136, 139-141 , suitably any one of SEQ ID NO: 136, 140, 141 , suitably any one of SEQ ID NO: 139-141 , suitably any one of SEQ ID NO: 140 and / or 141 , or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0303] Suitably, the oligonucleotide may target SEQ ID NO: 4. Suitably, the oligonucleotide may be an siRNA comprising or consisting a sequence selected from any one of SEQ ID NO: 140 and 141 , or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0304] Suitably, the oligonucleotide may target SEQ ID NO: 3 (ENSE00002440436). Suitably, the oligonucleotide may be an siRNA comprising or consisting of a sequence selected from any one of SEQ ID NO: 142-159, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably, the oligonucleotide may be an siRNA comprising or consisting of any one or more of those sequences that target any one or more of SEQ ID NO: 5-9 as mentioned hereinbelow (since SEQ ID NO: 5-9 are comprised within SEQ ID NO: 3).

[0305] Suitably, the oligonucleotide may target SEQ ID NO: 5. Suitably, the oligonucleotide may be an siRNA comprising or consisting of a sequence selected from any one of SEQ ID NO: 142- 146, 148-159, suitably any one of SEQ ID NO: 142-144, 146, 148-159, suitably any one of SEQ ID NO: 142-146, 148-156, 158-159, suitably any one of SEQ ID NO: 142-144, 146, 148- 156, 158-159, suitably any one of SEQ ID NO: 142-144, 146, 148-152, 154-156, 158-159, suitably any one of SEQ ID NO: 142-144, 146, 148-151 , 154-156, 158-159, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably, the oligonucleotide may be an siRNA comprising or consisting of any one or more of those sequences that target any one or more of SEQ ID NO: 6-9 as mentioned hereinbelow (since SEQ ID NO: 6-9 are comprised within SEQ ID NO: 5).

[0306] Suitably, the oligonucleotide may target SEQ ID NO: 6. Suitably, the oligonucleotide may be an siRNA comprising or consisting of a sequence selected from any one of SEQ ID NO: 142- 144, 148-159, suitably any one of SEQ ID NO: 142-144, 148-156, 158-159, suitably any one of SEQ ID NO: 142-144, 148-152, 154-156, 158-159, suitably any one of SEQ ID NO: 142- 144, 148-151 , 154-156, 158-159, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably, the oligonucleotide may be an siRNA comprising or consisting of any one or more of those sequences that target any one or more of SEQ ID NO: 7-9 as mentioned hereinbelow (since SEQ ID NO: 7-9 are comprised within SEQ ID NO: 6), suitably in combination with any one or more of SEQ ID NO: 152, 153, 157.

[0307] Suitably, the oligonucleotide may target SEQ ID NO: 7. Suitably may be an siRNA comprising or consisting of a sequence selected from any one of SEQ ID NO: 142, 143, 148-151 , or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably, the oligonucleotide may target SEQ ID NO: 8. Suitably may be an siRNA comprising or consisting of a sequence selected from any one of SEQ ID NO: 154-156, or a sequence having at least 50%, at least 60%, at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0308] Suitably, the oligonucleotide may target SEQ ID NO: 9. Suitably, the oligonucleotide may be an siRNA comprising or consisting of a sequence selected from any one of SEQ ID NO: 144, 158, 159, or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0309] Suitably the oligonucleotide may be an siRNA comprising or consisting of SEQ ID NO: 27 or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto.

[0310] The oligonucleotide may equally target any one or more of SEQ ID NO: 119, 120, 48-56, 121- 126.

[0311] Suitably, an oligonucleotide targeting SEQ ID NO: 119 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 7 and / or 8. Suitably, an oligonucleotide targeting SEQ ID NO: 120 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 8 and / or 9.

[0312] Suitably, an oligonucleotide targeting SEQ ID NO: 48 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1-3.

[0313] Suitably, an oligonucleotide targeting SEQ ID NO: 49 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 3.

[0314] Suitably, an oligonucleotide targeting SEQ ID NO: 50 may be an sRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 4 and / or 3. Suitably, an oligonucleotide targeting SEQ ID NO: 51 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 5.

[0315] Suitably, an oligonucleotide targeting SEQ ID NO: 52 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 4 and / or 5.

[0316] Suitably, an oligonucleotide targeting SEQ ID NO: 53 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 6.

[0317] Suitably, an oligonucleotide targeting SEQ ID NO: 54 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 4 and / or 6.

[0318] Suitably, an oligonucleotide targeting SEQ ID NO: 55 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 7.

[0319] Suitably, an oligonucleotide targeting SEQ ID NO: 56 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 4 and / or 7.

[0320] Suitably, an oligonucleotide targeting SEQ ID NO: 51 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 5.

[0321] Suitably, an oligonucleotide targeting SEQ ID NO: 121 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1, 2 and / or 5.

[0322] Suitably, an oligonucleotide targeting SEQ ID NO: 122 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1, 2 and / or 6.

[0323] Suitably, an oligonucleotide targeting SEQ ID NO: 123 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1, 2 and / or 7. Suitably, an oligonucleotide targeting SEQ ID NO: 124 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 1 , 2 and / or 119.

[0324] Suitably, an oligonucleotide targeting SEQ ID NO: 125 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 119.

[0325] Suitably, an oligonucleotide targeting SEQ ID NO: 126 may be an siRNA comprising or consisting of any one or more of those sequences that are comprised in oligonucleotides targeting SEQ ID NO: 2 and / or 120.

[0326] In one preferred embodiment, oligonucleotide may be an siRNA comprising or consisting of SEQ ID NO: 23-40.

[0327] The oligonucleotide may comprise a functional variant of any of the sequences mentioned above, suitably any of SEQ ID NO: 129-159. By “functional variant” is meant a variant which maintains it’s functionality in being able to reduce the level of MEG3 expression, suitably in by binding to MEG3. Functional variants may be determined by a skilled person using techniques described in the Examples. Functional variants may include modifications such as truncations or a small number of insertions, deletions and / or substitutions in a given sequence, suitably between 1-10 nucleotide modifications. Merely by way of an example, a functional variant of SEQ ID NO: 129 may include a single nucleotide substitution or for example a trinucleotide deletion in the sequence of SEQ ID NO: 129.

[0328] Reducing the level of MEG3 expression in a cell

[0329] Without wishing to be bound by theory, the level of MEG3 expression within the cell may be reduced through a number of different mechanisms. Reducing the level of MEG3 expression requires binding (i.e. hybridisation) of the oligonucleotide to MEG3 transcripts and / or the MEG3 gene, as described in more detail below.

[0330] Firstly, the level of MEG3 expression within the cell may be reduced by reducing or inhibiting transcription of the MEG3 gene. Accordingly, the oligonucleotide may reduce or inhibit, may be capable of reducing or inhibiting, may promote a reduction or inhibition of, or may be capable of promoting a reduction or inhibition of transcription of the MEG3 gene, suitably by binding to the MEG3 gene. Transcription of the MEG3 gene may be reduced or inhibited, through for example RNA-directed DNA methylation (RdDM), which is a widely studied mechanism of gene silencing that is mediated through the RNA Induced Transcriptional Silencing (RITS) complex, in what is commonly referred to as transcriptional gene silencing. Accordingly, the oligonucleotide may promote, or being capable or promoting, RdDM of the MEG3 gene i.e. RITS mediated reduction or inhibition of transcription of the MEG3 gene, suitably by binding to the MEG3 gene. Suitably, a reduction in the level of MEG3 expression in the cell may be mediated by the RdDM of the MEG3 gene i.e. RITS reduction or inhibition of MEG3 transcription. RdDM typically requires that the oligonucleotide comprises ribonucleotides, more typically that the oligonucleotide consists of ribonucleotides, yet more typically that the oligonucleotide is a short interfering RNA (siRNA) or short hairpin RNA (shRNA). Accordingly, an oligonucleotide capable of reducing or inhibiting transcription of the MEG3 gene, suitably by being capable of promoting RdDM of the MEG3 gene i.e. RITS mediated reduction or inhibition of transcription of the MEG3 gene, may comprise or consist of ribonucleotides (modified and / or unmodified), and suitably may be an siRNA or shRNA.

[0331] Secondly, the level of MEG3 expression in the cell may be reduced by reducing the level of MEG3 transcripts, such as by degrading MEG3 transcripts. Accordingly, the oligonucleotide may reduce, be capable of reducing, promote a reduction of, or be capable of promoting a reduction of the level of MEG3 transcripts in the cell, such as by degrading MEG3 transcripts in the cell, suitably by binding to MEG3 transcripts. The level of MEG3 transcripts may be reduced by the RNA Induced Silencing Complex (RISC) mediated degradation which is a widely studied mechanism of gene silencing that is commonly referred to as post- transcriptional gene silencing. Accordingly, the oligonucleotide may promote, or be capable or promoting, RISC degradation of MEG3 transcripts, suitably by binding to MEG3 transcripts. Suitably, a reduction in the level of MEG3 expression in the cell may be mediated by the RISC degradation of MEG3 transcripts. The RISC complex is guided to its target, for example to MEG3 transcripts, by an oligonucleotide that typically comprises ribonucleotides, more typically an oligonucleotide that consists of ribonucleotides, yet more typically an oligonucleotide which is processed from an siRNA or shRNA. Accordingly, an oligonucleotide capable of reducing the level of MEG3 transcripts in a cell, suitably by being capable of promoting RISC degradation of MEG3 transcripts, may comprise or consist of ribonucleotides (modified and / or unmodified), and suitably may be a siRNA or shRNA.

[0332] Additionally, or alternatively, the level of MEG3 transcripts may also be reduced by RNase H mediated degradation of MEG3 transcripts. Accordingly, the oligonucleotide may promote, or be capable of promoting, RNase H degradation of MEG3 transcripts, suitably by binding to MEG3 transcripts. Suitably, a reduction in the level of MEG3 expression in the cell may be mediated by RNase H degradation of MEG3 transcripts. RNase H recognises and cleaves DNA:RNA duplex structures, and so would degrade MEG3 transcripts that are bound to (i.e. hybridised to) antisense oligonucleotides comprising or consisting of deoxyribonucleotides. Accordingly, an oligonucleotide capable of reducing the level of MEG3 transcripts in the cell, such as by being capable of degrading MEG3 transcripts in the cell suitably by being capable of promoting RNase H degradation of MEG3 transcripts, may comprise or consist of deoxyribonucleotides (modified and / or unmodified), and suitably may be an ASO. Suitably said oligonucleotide is a mixmer ASO or a gapmer ASO, suitably comprising or consisting of deoxyribonucleotides (modified and / or unmodified). Suitably said oligonucleotide is a gapmer ASO wherein the central portion of the gapmer ASO comprises or consists of deoxyribonucleotides (modified and / or unmodified), suitably the central portion of the gapmer ASO consists of deoxyribonucleotides (modified and / or unmodified).

[0333] Thirdly, the level of MEG3 expression in the cell may be reduced by reducing, inhibiting or repressing the translation of MEG3 transcripts. Accordingly, the oligonucleotide may reduce, inhibit or repress; be capable of reducing, inhibiting or repressing; promote a reduction, inhibition or repression of; or be capable of promoting a reduction, inhibition or repression of the translation of MEG3 transcripts, suitably by translation inhibition or translational repression of MEG3 transcripts, suitably by binding to MEG3 transcripts. Translation of MEG3 transcripts may be reduced, inhibited or repressed by the oligonucleotide binding to MEG3 transcripts and thereby blocking access to (i.e. sterically occluding) the molecules required for translation to take place (the translation machinery). Accordingly, the oligonucleotide may sterically occlude access to MEG3 transcripts of molecules required for translation, suitably of the translation machinery. Steric occlusion typically requires that the oligonucleotide is able to bind to it’s target. Accordingly, an oligonucleotide capable of reducing, inhibiting or repressing the translation of MEG3 transcripts, suitably by steric occlusion, may be able to bind to MEG3 transcripts, and suitably therefore may comprise or consist of deoxyribonucleotides (modified and / or unmodified) and / or ribonucleotides (modified and / or unmodified).

[0334] Other mechanisms may exist whereby the level of MEG3 expression in the cell may be reduced by the oligonucleotide, including any other mechanism known in the art.

[0335] The oligonucleotide of the invention may reduce the level of, or may reduce, inhibit or repress the translation of, one or more MEG3 transcript isoforms. The oligonucleotide of the invention may reduce the level of, or may reduce, inhibit or repress the translation of, a plurality of MEG3 transcript isoforms. Where the oligonucleotide binds to the MEG3 gene leading to reduced or inhibited transcription of the MEG3 gene, the level of all transcript isoforms of MEG3 may be reduced. Where the oligonucleotide binds to MEG3 transcripts, the level of those transcript isoforms to which the oligonucleotide may bind are reduced. The oligonucleotide may reduce or be capable of reducing the level of MEG3 expression within a cell by 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%,

[0336] 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%,

[0337] 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%,

[0338] 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%,

[0339] 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,

[0340] 97%, 98%, or 99% or more relative to the level of MEG3 expression in a reference cell. Suitably wherein a reference is the same cell before the oligonucleotide is applied, or a control cell such as a cell of the same type which is not treated with the oligonucleotide, suitably wherein a control cell may be a healthy cell or a cell from a healthy subject.

[0341] The oligonucleotide may reduce or be capable of reducing the level of MEG3 expression to a level that is equivalent to the expression of MEG3 in a control cell. Suitably therefore the oligonucleotide may partially silence the MEG3 gene within a cell. In one embodiment, the oligonucleotide effectively silences the MEG3 gene within a cell.

[0342] Suitably the level of MEG3 expression refers to the level of MEG3 gene expression, suitably within a given cell. Suitably which may be determined by the level of MEG3 transcripts present in a cell, which my suitably be measured by conventional techniques such as qPCR or RNA- seq. Suitably the oligonucleotide may reduce or be capable of reducing the level of MEG3 expression in any cell, suitably in any cell which typically expresses the MEG3 gene. Suitably within muscle cells, cardiac cells, cardiomyocyte cells, nerve cells, brain cells, lung cells, pancreatic cells, eye cells such as photoreceptor cells, cartilage cells such as chondrocytes, or bone marrow cells such as blood stem cells including for example myeloid progenitor cells and haematopoietic progenitor cells.

[0343] Suitably, an oligonucleotide targeting the region SEQ ID NO: 1 (or SEQ ID NO: 48, 121-124) may reduce or be capable of reducing the level of MEG3 expression within the cell by 20% or more, 24% or more, 30% or more, 33% or more, 41% or more, 50% or more, 53% or more, 55% or more, 57% or more, 60% or more, 65% or more, 70% or more, 75% or more, 80% or more, 85% or more, 90% or more, or 95% or more. In one embodiment, by 57% or more. In one embodiment, by 65% or more. Suitably compared to the expression in a reference cell. Suitably, the region of SEQ ID NO: 1 (or SEQ ID NO: 48, 121-124), or a target sequence comprised in region SEQ ID NO: 1 (or SEQ ID NO: 48, 121-124), when targeted using an oligonucleotide, may lead to these levels of reduction in MEG3 expression. Suitably, an oligonucleotide targeting the regions SEQ ID NO: 2 or SEQ ID NO: 4 (or any one of SEQ ID NO: 48-56, 121-126) may reduce or be capable of reducing the level of MEG3 expression within the cell by 10% or more, 15% or more, 17% or more, 20% or more, 26% or more, 30% or more, 40% or more, 50% or more, 60% or more, 61% or more, 65% or more, 70% or more, 75% or more, 76% or more, 78% or more, 80% or more, 84% or more, 90% or more, or 95% or more. In one embodiment, by 61% or more. In one embodiment, by 76% or more. In one embodiment, by 78% or more. Suitably compared to the expression in a reference cell. Suitably, the region SEQ ID NO: 2 or SEQ ID NO: 4 (or any one of SEQ ID NO: 48-56, 121-126), or a target sequence comprised in region SEQ ID NO: 2 or SEQ ID NO: 4 (or any one of SEQ ID NO: 48-56, 121-126), when targeted using an oligonucleotide, may lead to these levels of reduction in MEG3 expression.

[0344] Suitably, an oligonucleotide targeting the regions SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8 or SEQ ID NO: 9 (or any one of SEQ ID NO: 48-56, 119-126) may reduce or be capable of reducing the level of MEG3 expression within the cell by 40% or more, 45% or more, 46% or more, 50% or more, 55% or more, 60% or more, 65% or more, 66% or more, 70% or more, 72% or more, 73% or more, 75% or more, 80% or more, 81% or more, 82% or more, 83% or more, 84% or more, 85% or more, 86% or more, 87% or more, 88% or more, 89% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more. In one embodiment, by 56% or more. In one embodiment, by 66% or more. In one embodiment, by 72% or more. In one embodiment, by 73% or more. In one embodiment, by 79% or more. In one embodiment, by 81% or more. In one embodiment, by 84% or more. In one embodiment, by 87% or more. In one embodiment, by 90% or more. Suitably compared to the expression in a reference cell. Suitably, the region SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8 or SEQ ID NO: 9 (or any one of SEQ ID NO: 48-56, 119-126), or a target sequence comprised in region SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8 or SEQ ID NO: 9 (or any one of SEQ ID NO: 48-56, 119-126), when targeted using an oligonucleotide, may lead to these levels of reduction in MEG3 expression.

[0345] Suitably, substantially all nucleotides targeting SEQ ID NO: 4 (or any one of SEQ ID NO: 48- 56, 121-126) may reduce or be capable of reducing the level of MEG3 expression within the cell by 70% or more, 75% or more, 80% or more, 81 % or more or 82% or more. Suitably compared to the expression in a reference cell.

[0346] Suitably, substantially all nucleotides targeting SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8 or SEQ ID NO: 9 (or any one of SEQ ID NO: 48-56, 119-126) may reduce or be capable of reducing the level of MEG3 expression within the cell by 60% or more, 66% or more, 70% or more, 72% or more, 73% or more, 74% or more, 75% or more, 76% or more, 77% or more, 78% or more, 79% or more, 80% or more, 81% or more, 82% or more, 83% or more, 84% or more, 85% or more, 86% or more, 87% or more, 88% or more, 89% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more. Suitably compared to the expression in a reference cell.

[0347] Vectors

[0348] The invention also provides related products comprising or encoding the oligonucleotide. For instance, the oligonucleotide of the invention may be encoded by a nucleic acid, which in turn may be comprised in a vector. “Encoded” is used herein to also encompass the oligonucleotide being comprised in the nucleic acid. Accordingly, the invention provides a vector comprising a nucleic acid encoding the oligonucleotide of the invention. The vector may be referred to herein as the vector of the invention.

[0349] “Vector” as used herein refers to a nucleotide vector, which may be linear or circular. Linear vectors may include linear DNA or RNA molecules. Circular vectors may circular DNA or RNA molecules including plasmids such as expression plasmids.

[0350] Suitably the vector may be a self-replicating vector such as a self-replicating plasmid, suitably a self-replicating DNA or RNA plasmid. Suitably a DNA or RNA vector may not express the oligonucleotide but instead may comprise a nucleic acid which itself is the oligonucleotide. For example, the vector may be a self-replicating RNA vector comprising an RNA ASO, a siRNA or an shRNA of the invention. For example, the vector may be a self-replicating DNA vector comprising a DNA ASO of the invention.

[0351] Whether the vector is linear or circular, the vector may be an expression vector such as an expression plasmid, suitably one which is capable of expressing the oligonucleotide.

[0352] In an expression vector, the nucleic acid encoding the oligonucleotide is typically operably linked to a control or regulatory element which is capable of providing for the expression of the nucleic acid by a host cell into which the expression vector may be introduced. Such expression vectors can be used to express the construct.

[0353] Suitable regulatory elements may include promoters, enhancers, UTRs, introns, etc. Typically an expression vector at least comprises a promoter which is suitable to promote expression of the nucleic acid encoding the oligonucleotide of the invention. Suitably the promoter may be an inducible promoter, which may be induced upon contact with an inducer, suitably the promoter may be induced after the vector has been introduced to a cell or administered to a subject to promote expression of the nucleic acid in the cell or subject.

[0354] The term “operably linked” refers to a juxtaposition wherein the components described are in a relationship permitting them to function in their intended manner. A control element “operably linked” to a coding sequence is ligated in such a way that expression of the coding sequence is achieved under conditions compatible with the control element.

[0355] Numerous expression vectors may be employed. Of course, any expression vector that is capable of eliciting expression in bacterial or eukaryotic cells may be used. Examples of suitable vectors include, but are not limited to plasmids pcDNA3, pHCMV / Zeo, pCR3.1 , pEF 1 / His, pIND / GS, pRc / HCMV2, pSV40 / Zeo2, pTRACER-HCMV, pUB6 / V5-His, pVAXI, and pZeoSV2 (available from Invitrogen, San Diego, Calif.), and plasmid pCI (available from Promega, Madison, Wis.). Suitable vectors may also include those used for gene therapy such as AAV vectors or lentiviral vectors, suitably for expressing the oligonucleotide in a subject.

[0356] The construction of vectors comprising a nucleic acid encoding the oligonucleotide is typically considered routine for a person skilled in the art, and will follow conventional molecular biological techniques such as those described in, for example, Molecular Cloning: A Laboratory Manual, second edition (Sambrook, et al., 1989) Cold Spring Harbor Press, or Gene Transfer Vectors for Mammalian Cells (J. M. Miller and M. P. Calos, eds., 1987).

[0357] The expression vector may be introduced into a suitable host cell, which may be a host cell in a subject to which the vector has been administered. Thus, a nucleic acid sequence comprising or encoding the oligonucleotide may be inserted into an expression vector, said expression vector may be introduced into a compatible host cell for example by administering it to a subject, and conditions may be present or induced which bring about expression of the nucleic acid comprising or encoding the oligonucleotide.

[0358] Bacterial host cells may be used for propagation of the vector or the oligonucleotide encoded by the nucleic acid. Accordingly therefore, there is provided a host cell, suitably a bacterial or eukaryotic host cell, comprising the vector described herein.

[0359] The vector may encode any form of the oligonucleotide disclosed herein that is capable of being encoded by a nucleic acid and expressed into the oligonucleotide. Vectors ASOs

[0360] Suitably, the vector may comprise a nucleic acid encoding an ASO, suitably wherein the ASO consists of deoxyribonucleotides or ribonucleotides. DNA ASOs may be produced from a DNA vector by DNA-dependent DNA polymerases. RNA ASOs may be produced from a DNA vector by DNA-dependent RNA polymerases. DNA ASOs may be produced from an RNA vector by reverse transcriptases. RNA ASOs may be produced from an RNA vector by RNA-dependent RNA polymerases. Where the ASO consists of ribonucleotides, expression of the nucleic acid may be driven by a promoter, which may or may not lead to polyadenylation of the ASO RNA transcript.

[0361] ASOs as the name suggests are typically encoded in an antisense orientation such that the RNA oligonucleotide transcribed from the nucleic acid (the RNA ASO) will be of an antisense orientation to the target sequence of interest (i.e. such that the expressed ASO is able to bind or hybridise to MEG3 transcripts).

[0362] Vectors encoding siRNAs

[0363] Suitably, the vector may comprise a nucleic acid encoding an siRNA. An siRNA may be encoded by a nucleic in an RNA vector. Suitably, the two strands of the siRNA may be comprised in two nucleic acids present on the same or two different RNA vectors.

[0364] An siRNA may be encoded by a nucleic acid in a DNA vector, suitably by two nucleic acids, each encoding one of the two complementary strands of the siRNA. Once the two RNAs are expressed from the two nucleic acids they may then hybridise to form an siRNA.

[0365] Accordingly, the invention provides a vector comprising at least one nucleic acid encoding the oligonucleotide of the invention. Accordingly, the invention provides a vector comprising two nucleic acid encoding the oligonucleotide of the invention, suitably wherein the oligonucleotide is an siRNA.

[0366] Vectors encoding shRNAs

[0367] Vectors are commonly used to express shRNAs. Accordingly, the skilled person will be aware how to construct vectors comprising a nucleic acid encoding shRNAs of the invention. Widely used promoters for expressing shRNAs from expression plasmids include inducible promoters, RNA polymerase II promoters such as CMV and EF1 , and RNA polymerase III promoters such as U3, U6, H1 or 7SK promoters for example. Suitably the vector may comprise an shRNA sequence designed to target any region or target sequence described herein according to shRNA designs known in the art or a sequence having at least 50%, at least 60%, at least 70% identity thereto, optionally at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% identity thereto. Suitably the vector may comprise one or more of said sequences, which may suitably target and bind to different target sequences.

[0368] Virus particles

[0369] The oligonucleotide or vector of the invention, or indeed a nucleic acid comprising or encoding the oligonucleotide of the invention, may be comprised in a virus particle. Accordingly, the invention provides a virus particle comprising the oligonucleotide or vector of the invention, or a nucleic acid encoding the oligonucleotide of the invention. For example ASOs, siRNAs and shRNAs may all be comprised in a virus particle, encoded by a nucleic acid comprised in a virus particle, or encoded by a vector comprised in a virus particle. The virus particle may be referred to herein as the virus particle of the invention.

[0370] The virus particle may be one that can be used in gene therapy, i.e. a viral particle that comprises all the required functional elements to express the nucleic acid (which encodes the oligonucleotide) in a host cell after administration as a therapeutic. Suitably in such embodiments, the host cell may be for example a muscle cell, cardiac cell, cardiomyocyte cell, nerve cell, brain cell, lung cell, pancreatic cell, eye cell such as a photoreceptor cell, cartilage cells such as chondrocytes, or bone marrow cell such as a blood stem cell including for example myeloid progenitor cells and haematopoietic progenitor cells. The virus particle may comprise a viral genome which comprises an expression cassette. The expression cassette may comprise or encode the oligonucleotide operably linked to a promoter. The promoter may be specific for certain cell types, such as muscle cells, cardiac cells, cardiomyocyte cells, nerve cells, brain cells, lung cells, pancreatic cells, eye cells (such as photoreceptor cells), cartilage cells (such as chondrocytes), or bone marrow cells such as blood stem cells including for example myeloid progenitor cells and haematopoietic progenitor cells.

[0371] Virus particles may comprise phages, viruses, virus-like particles (VLPs), retroviruses, and any other virus particles or virus-related particles known in the art for delivering nucleic acids to cells. Suitable virus particles include or may be derived from for example a parvovirus, a retrovirus, a lentivirus, or a herpes simplex virus. The parvovirus may be an adeno-associated virus (AAV). Suitably the virus particle may be recombinant adeno-associated viral (AAV) vector or a lentiviral vector. Suitably, the virus particles may be an AAV virus particles (i.e. viral vector).

[0372] Conjugates

[0373] The invention also provides related products comprising or encoding the oligonucleotide or indeed the vector. Accordingly, the invention provides a conjugate comprising the oligonucleotide or vector of the invention covalently linked to a delivery group. For example, ASOs, siRNAs and shRNAs may be comprised in a conjugate, more typically ASOs and siRNAs. The conjugate may be referred to herein as the conjugate of the invention.

[0374] Suitably a conjugate of the invention may comprise any delivery group capable of aiding delivery or uptake of the oligonucleotide into a target cell. Suitable such delivery groups may include: antibodies, peptides, small molecule chemicals, polymers, lipids, or the like. Suitably therefore, the oligonucleotide may be conjugated to a delivery group.

[0375] Suitably the delivery group may be a peptide. Suitably, the peptide may be selected from viral proteins such as VP22 (derived from herpes virus tegument protein), snake venom protein such as CyLOP-1 (derived from crotamin), cell adhesion glycoproteins such as pVEC (derived from murine vascular endothelial-cadherin protein), Penetratin (Antennapedia homeodomain), Tat (human immunodeficiency virus transactivating regulatory protein), cell penetrating peptide (CPP), or reverse Tat, for example. In some embodiments, the oligonucleotide is conjugated to a delivery group wherein the delivery group is a cell penetrating peptide (CPP), suitably to enhance therapeutic uptake.

[0376] Cell penetrating peptides (CPPs) are typically short peptides primarily composed of basic residues and are typically rich in lysine and arginine residues. CPPs are believed to interact with negatively charged cell surface molecules through these positively charged amino acid residues, thereby help the peptides naturally cross the cell membrane commonly by endocytosis.

[0377] Suitable CPPs are known in the art. The use of arginine-rich peptide carriers may be particularly useful. Certain arginine based peptide carriers have been shown to be highly effective at delivery of antisense compounds into primary cells including muscle cells (Marshall, Oda et al. 2007; Jearawiriyapaisarn, Moulton et al. 2008; Wu, Moulton et al. 2008). Furthermore, compared to other peptides, the arginine peptide carriers when conjugated to an antisense oligonucleotide (or indeed an siRNA for example), demonstrate an enhanced ability to alter transcript levels (Marshall, Oda et al. 2007). Suitably, the cell penetrating peptide may be selected from those peptides described in WO2015075747, WO2013030569, WO2009147368, US20120289457, or US20160237426, for example.

[0378] By conjugated it is meant that the oligonucleotide is covalently linked to the delivery group. Suitably therefore, conjugate comprises the oligonucleotide covalently linked to a delivery group. Suitably, conjugation of the delivery group to the oligonucleotide may be at any position suitable for forming a covalent bond between the delivery group and the oligonucleotides or between a linker and the oligonucleotide. For example, conjugation of a delivery group may be at the 3' end of the oligonucleotide. Alternatively, conjugation of a delivery group to the oligonucleotide may be at the 5' end of the oligonucleotide. Alternatively, a delivery group may be conjugated to the oligonucleotide through any of the internucleoside linkage groups. Alternatively, multiple delivery groups may be conjugated at the 5’ end, 3’ end and / or at any of the internucleoside linkage groups.

[0379] Suitably, the delivery group may be covalently coupled at its N-terminal or C-terminal residue to the 3' or 5' end of the oligonucleotide. Suitably, the delivery group may be coupled at its C- terminal residue to the 5' end of the oligonucleotide. Suitably, the delivery group may be coupled at its N-terminal residue to the 5' end of the oligonucleotide. Suitably, the delivery group may be coupled at its C-terminal residue to the 3' end of the oligonucleotide. Suitably, the delivery group may be coupled at its N-terminal residue to the 3' end of the oligonucleotide.

[0380] Optionally, where the oligonucleotide comprises phosphorus-containing internucleoside linkages, and the delivery group is a peptide, the peptide may be conjugated to the oligonucleotide via a covalent bond to the phosphorous, suitably the phosphorous of the terminal linkage group.

[0381] Alternatively, when the delivery group is a peptide, and the oligonucleotide is a morpholino, the peptide may be conjugated to the nitrogen atom of the 3' terminal morpholino group of the oligomer for example.

[0382] Optionally, the delivery group may be conjugated to the oligonucleotide via a linker. Optionally, the linker may be any linker known in the art, suitably the linker may comprise one or more of: an optionally substituted piperazinyl moiety, a beta alanine, glycine, serine, threonine, a hydrocarbon chain, proline, 6-aminohexanoic acid residue, lysine and / or arginine in any combination. A typically linker may be a short chain of glycine and serine residues, for example

[0383] Alternatively, the delivery group may be conjugated directly to the oligonucleotide without a linker. Suitably the conjugate may comprise or further comprise a homing moiety i.e. tissue targeting moiety. Suitably, the homing moiety is selective for a selected tissue such as a mammalian tissue, i.e., the same tissue being targeted by the oligonucleotide. Suitably, the homing moiety may be selective for muscle tissue, cardiac tissue, heart muscle tissue, nervous tissue, lung tissue, pancreatic tissue, eye tissue, cartilage tissue, eye tissue, bone marrow tissue or any tissue of the central nervous system (CNS) including brain and spinal cord tissue.

[0384] Suitably, the homing moiety may be a homing peptide. Suitable homing peptides are disclosed in 'Effective Dystrophin Restoration by a Novel Muscle-Homing Peptide-Morpholino Conjugate in Dystrophin-Deficient mdx Mice' Gao et al. Mol Ther. 2014 Jul; 22(7): 1333-1341 , for example.

[0385] Suitably, where the conjugate comprises both a delivery peptide and a homing peptide, the delivery peptide and the homing peptide may be formed as a chimeric fusion protein. Suitably, the conjugate may comprise a chimeric peptide formed from a cell penetrating peptide and for example a nervous tissue-specific homing peptide (or for example a muscle tissue-, cardiac tissue-, heart muscle tissue-, nervous tissue-, brain tissue-, spinal cord tissue-, lung tissue-, pancreatic tissue-, cartilage tissue-, eye tissue- or bone marrow tissue-specific homing peptide). Optionally, the conjugate may be of the form: delivery peptide-homing peptideoligonucleotide, or of the form: homing peptide-carrier peptide-oligonucleotide, or of the form: oligonucleotide-delivery peptide-homing peptide, or of the form: oligonucleotide-homing peptide-delivery peptide. Optionally such forms of the conjugate may comprise linkers located between each element.

[0386] Suitably, the oligonucleotide may be conjugated to a delivery group that enhances the solubility of the oligonucleotide. Suitably the solubility in an aqueous medium. Suitably, a delivery group that enhances solubility may for example be conjugated to the oligonucleotide in addition to a delivery group operable to transport the oligonucleotide. Suitable delivery groups that enhance the solubility of a oligonucleotide are known in the art and include polymers, such as polyethylene glycol, or triethylene glycol.

[0387] Suitably, the delivery group that enhances solubility and the delivery group that transports the oligonucleotide may be formed as a chimeric fusion protein.

[0388] Suitably, the delivery group has the capability of inducing cell penetration of the oligonucleotide within at least 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of cells of a given cell culture population. Suitably, the delivery group has the capability of inducing cell penetration of the oligonucleotide within at least 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of muscle cells, cardiac cells, cardiomyocyte cells, nerve cells, brain cells, lung cells, pancreatic cells, eye cells, cartilage cells (such as chondrocytes) or bone marrow cells such as blood stem cells including for example myeloid progenitor cells and haematopoietic progenitor cells.

[0389] Delivery particles

[0390] The oligonucleotide, vector, or conjugate of the invention may be comprised within a particle, suitable within a delivery particle. The invention therefore provides a delivery particle comprising the oligonucleotide, vector, or conjugate of the invention. The delivery particle may be referred to herein as the delivery particle of the invention.

[0391] Suitably the delivery particle may be capable of aiding delivery or uptake of the oligonucleotide into a target cell. Suitable delivery particles may include: nanoparticles including polymeric nanoparticles, nanocarriers including supramolecular nanocarriers, vesicles or the like. Suitably the delivery particle may be a nanoparticle or a vesicle, suitably a nanoparticle.

[0392] Supramolecular nanocarriers may include cationic polymer complexes and various polymeric compounds. Complexation of nucleic acids with various polycations is another approach for intracellular delivery. This includes use of PEGylated polycations, polyethyleneamine (PEI) complexes, cationic block co-polymers, and dendrimers. Several cationic nanocarriers, including PEI and polyamidoamine dendrimers help to release contents from endosomes.

[0393] Other approaches include use of microspheres including biodegradable microspheres, dendrimers, biodegradable polymers, prodrugs, inorganic colloids such as sulphur or iron, antibodies, implants including biodegradable implants and osmotically controlled implants, lipid nanoparticles, emulsions, oily solutions, aqueous solutions, biodegradable polymers, poly(lactide-coglycolic acid), poly(lactic acid), liquid depot, polymer micelles, quantum dots and lipoplexes. In some embodiments, an oligonucleotide is conjugated to another molecule which is known in the art to act as a carrier.

[0394] Vesicles may include extracellular vesicles, liposomes, lipid vesicles, lipid nanoparticles (LNPs), exosomes or the like. Vesicles such as LPNs for example are typically capable of being endocytosed by cells, and may be decorated with targeting moieties to target these to specific tissues of interest.

[0395] In some embodiments, the oligonucleotide, conjugate, or vector of the invention is comprised within a nanocarrier. In some embodiments, the oligonucleotide, conjugate, or vector of the invention is comprised within a vesicle, suitably a liposome or lipid nanoparticle (LNP).

[0396] Suitably, the delivery particle has the capability of inducing cell penetration of the oligonucleotide within at least 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of cells of a given cell culture population. Suitably, the delivery particle has the capability of inducing cell penetration of the oligonucleotide within at least 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of muscle cells, cardiac cells, cardiomyocyte cells, nerve cells, brain cells, lung cells, pancreatic cells, cartilage cells (such as chrondrocytes), eye cells or bone marrow cells such as blood stem cells including for example myeloid progenitor cells and haematopoietic progenitor cells.

[0397] Use in research

[0398] The oligonucleotide, vector, virus particle, conjugate, or delivery particle of the invention, or a pharmaceutical composition of any of these described elsewhere herein, may be used in research. Accordingly, the invention provides the oligonucleotide, vector, virus particle, conjugate, or the delivery particle of the invention, or a pharmaceutical composition thereof, for use in research.

[0399] Pharmaceutical compositions

[0400] The oligonucleotide, vector, virus particle, conjugate or delivery particle of the invention may be formulated in any suitable manner. Accordingly, the invention provides a pharmaceutical composition comprising the oligonucleotide, vector, virus particle, conjugate or the delivery particle of the invention, suitably a pharmaceutical composition comprising any combination thereof. The pharmaceutical composition may be referred herein to as the pharmaceutical composition of the invention.

[0401] A "pharmaceutical composition" refers to a preparation which the one or more active ingredients (i.e. the oligonucleotide, vector, virus particle, conjugate and / or delivery particle) is in a form that permits their biological activity to be effective, and which contains no additional components which are unacceptably toxic to a subject. A pharmaceutical composition may not be without side-effects however.

[0402] Optionally, the oligonucleotide may be present in the pharmaceutical composition as a physiologically tolerated salt. Suitably, physiologically tolerated salts retain the desired biological activity of the oligonucleotide and do not impart undesired toxicological effects. For oligonucleotides, suitable examples of pharmaceutically acceptable salts include (a) salts formed with cations such as sodium, potassium, ammonium, magnesium, calcium, polyamines such as spermine and spermidine, etc; (b) acid addition salts formed with inorganic acids, for example hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid and the like; (c) salts formed with organic acids such as, for example, acetic acid, oxalic acid, tartaric acid, succinic acid, maleic acid, fumaric acid, gluconic acid, citric acid, malic acid, ascorbic acid, benzoic acid, tannic acid, palmitic acid, alginic acid, polyglutamic acid, naphthalenesulfonic acid, methanesulfonic acid, p-toluenesulfonic acid, naphthalenedisulfonic acid, polygalacturonic acid, and the like; and (d) salts formed from elemental anions such as chlorine, bromine, and iodine.

[0403] Optionally the oligonucleotides within the composition may be chirally-controlled. Suitably therefore the composition may be regarded as a chirally controlled oligonucleotide composition.

[0404] In some embodiments, any of the pharmaceutical compositions according to the invention may further comprise a pharmaceutically acceptable excipient such as a carrier, diluent or solvent, buffer, stabiliser, additive, vehicle, antioxidant, adjuvant, bulking agent, preservative, encapsulating agent or other materials well known to those skilled in the art. Such materials should be non-toxic and should not interfere with the efficacy of the active ingredient.

[0405] A "pharmaceutically acceptable excipient" refers to an ingredient in a pharmaceutical formulation other than the active ingredient, which is not unreasonably toxic to a subject. Suitable pharmaceutically acceptable excipients are well known in the art. Each excipient must be "acceptable" in the sense of being compatible with the other components of the composition e.g. the oligonucleotide, and must not be injurious or toxic to the individual. Suitable pharmaceutically acceptable excipients are described, for example, in Remington's Pharmaceutical Sciences: The Science and Practice of Pharmacy, 20th ed, 2000, pub. Lippincott, Williams & Wilkins.

[0406] Suitable carriers may include, for example, water, saline, ethanol, glycerol, propylene glycol, polyethylene glycol, mineral oil, vegetable oils, gelatin, lactose, starch, cellulose, microcrystalline cellulose, dicalcium phosphate, calcium carbonate, sorbitol, mannitol, sucrose, dextrose, benzyl alcohol, stearic acid, magnesium stearate, sodium chloride, sodium citrate, citric acid, sodium phosphate, potassium phosphate, triglycerides, polysorbates (such as polysorbate 80), polyvinylpyrrolidone (PVP), carboxymethylcellulose, methylcellulose, hydroxypropyl methylcellulose, chitosan, lecithin, albumin and / or any other carrier known in the art. A mixture of two or more carriers may be used. Suitable diluents or solvents solvents may include, for example, water, ethanol, glycerin, propylene glycol, polyethylene glycol, benzyl alcohol, isopropanol, dimethyl sulfoxide (DMSO), acetone, ethyl acetate, methanol, butanol, isopropyl myristate, triacetin, N-methyl-2- pyrrolidone (NMP), diethylene glycol monoethyl ether, mineral oil, vegetable oils, and / or any other solvent known in the art. A mixture of two or more solvents may be used.

[0407] Suitable buffers may include, for example, tris, HEPES, MOPS, MES, acetate, phosphate, citrate, bicarbonate, succinate, borate, glycine, histidine, lactate, maleate, tartrate, carbonate, other organic acids, and / or any other buffer known in the art. A mixture of two or more buffers may be used. The buffer or the mixture of two or more buffer may typically be present in an amount of about 0.001% to about 10% by weight of the pharmaceutical composition. A mixture of two or more buffers may be used.

[0408] Suitable antioxidants may include, for example, ascorbic acid, tocopherol, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), propyl gallate, sodium metabisulfite, sodium bisulfite, sodium thiosulfate, citric acid, tartaric acid, glutathione, methionine, alpha- lipoic acid, cysteine, EDTA (ethylenediaminetetraacetic acid), ferulic acid, resveratrol, quercetin, carotenoids, uric acid, selenium, zinc, manganese, coenzyme Q10, lycopene, betacarotene, lutein, zeaxanthin, curcumin, flavonoids, polyphenols and / or any other antioxidant known in the art. A mixture of two or more antioxidants may be used.

[0409] Suitable bulking agents may include, for example, lactose, sucrose, mannitol, sorbitol, microcrystalline cellulose, dicalcium phosphate, calcium carbonate, starch, pregelatinized starch, maltodextrin, polyethylene glycol, polyvinylpyrrolidone (PVP), cellulose derivatives (such as hydroxypropyl cellulose and methylcellulose), sodium chloride, talc, silica and / or any other known bulking agent known in the art. A mixture of two or more bulking agents may be used.

[0410] Suitable preservatives may include various antibacterial and antifungal agents, for example, octadecyldimethylbenzyl ammonium chloride, benzalkonium chloride, benzethonium chloride, phenol, sorbic acid, butyl alcohol, benzyl alcohol, chlorobutanol, parabens such as methyl paraben and propyl paraben, catechol, resorcinol, cyclohexanol, 3-pentanol, m-cresol, sodium benzoate and / or any other preservatives known in the art. A mixture of two or more preservatives may be used. The preservative or mixture of preservative are typically present in an amount of about 0.0001 to about 2% by weight of the total pharmaceutical composition. A mixture of two or more preservatives may be used. Suitable encapsulating agents include, for example gelatin, hypromellose (hydroxypropyl methylcellulose), cellulose acetate phthalate, Eudragit polymers, ethyl cellulose, polyvinyl alcohol, alginate, chitosan, zein, acacia, pectin, carrageenan, starch, gum Arabic and / or any other encapsulating agent known in the art. A mixture of two or more encapsulating agents may be used.

[0411] Suitable additives may include, for example, proteins such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, valine, selenocysteine, pyrrolysine; monosaccharides, disaccharides and other carbohydrates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counter-ions such as sodium; metal complexes (e.g. Zn-protein complexes); and / or non-ionic surfactants such as polyethylene glycol (PEG). A mixture of two or more additives may be used.

[0412] Suitable additives may also include one or more active ingredients useful for the particular indication, disease, or condition being treated with the pharmaceutical composition, preferably those with activities complementary to the oligonucleotide, vector, virus particle, conjugate and / or delivery particle of the invention, where the respective activities do not adversely affect one another. Such active ingredients are suitably present in combination in amounts that are effective for the purpose intended.

[0413] The pharmaceutical carrier or diluent may be, for example, an isotonic solution. The precise nature of the carrier or other material may depend on the route of administration, e.g. oral, intravenous, cutaneous or subcutaneous, nasal, intramuscular and intraperitoneal routes. Examples of suitable compositions and methods of administration are provided in Esseku and Adeyeye (2011) and Van den Mooter G. (2006). For example, solid oral forms may contain, together with the active substance, diluents, e.g. lactose, dextrose, saccharose, cellulose, corn starch or potato starch; lubricants, e.g. silica, talc, stearic acid, magnesium or calcium stearate, and / or polyethylene glycols; binding agents; e.g. starches, gum arabic, gelatin, methylcellulose, carboxymethylcellulose or polyvinyl pyrrolidone; disaggregating agents, e.g. starch, alginic acid, alginates or sodium starch glycolate; effervescing mixtures; dyestuffs; sweeteners; wetting agents, such as lecithin, polysorbates, laurylsulphates; and, in general, non-toxic and pharmacologically inactive substances used in pharmaceutical formulations. Such pharmaceutical preparations may be manufactured in known manner, for example, by means of mixing, granulating, tableting, sugar-coating, or film-coating processes. The pharmaceutical composition may, for example, be in a form of a capsule, tablet, pill, liquid formulation, dispersant, granule, microgranule, film-coated tablet, pellet, lozenge, sublingual formulation, peptizer, buccal tablet, paste, syrup, suspension, elixir, emulsion, coating formulation, ointment, plaster, cataplasm, transdermal patch, lotion, inhalant, aerosol, eye drop, injection, or suppository.

[0414] Oral formulations include such normally employed excipients as, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate, and the like. These compositions take the form of solutions, suspensions, tablets, pills, capsules, sustained release formulations or powders and contain 10% to 95% of active ingredient, preferably 25% to 70%. Where the pharmaceutical composition is lyophilised, the lyophilised material may be reconstituted prior to administration, e.g. a suspension. Reconstitution is suitably effected in buffer.

[0415] Capsules, tablets and pills for oral administration to an individual may be provided with an enteric coating comprising, for example, Eudragit“S”, Eudragit“L”, cellulose acetate, cellulose acetate phthalate or hydroxypropylmethyl cellulose.

[0416] Liquid dispersions for oral administration may be syrups, emulsions or suspensions. The syrups may contain as carriers, for example, saccharose or saccharose with glycerine and / or mannitol and / or sorbitol.

[0417] Suspensions and emulsions may contain as carrier, for example a natural gum, agar, sodium alginate, pectin, methylcellulose, carboxymethylcellulose, or polyvinyl alcohol. The suspensions or solutions for intramuscular injections may contain, together with the active substance, a pharmaceutically acceptable carrier, e.g. sterile water, olive oil, ethyl oleate, glycols, e.g. propylene glycol, and if desired, a suitable amount of lidocaine hydrochloride.

[0418] Solutions for intravenous administration or infusion may contain as carrier, for example, sterile water or suitably they may be in the form of sterile, aqueous, isotonic saline solutions.

[0419] For suppositories, traditional binders and carriers may include, for example, polyalkylene glycols or triglycerides; such suppositories may be formed from mixtures containing the active ingredient in the range of 0.5% to 10%, preferably 1% to 2%.

[0420] In some embodiments, the pharmaceutical composition may be formulated for intravenous, arterial, intradermal, intramuscular, intrathecal, intraparenchymal, intraperitoneal, subcutaneous, sublingual or oral administration. In one embodiment, the pharmaceutical composition is formulated for intravenous or intrathecal administration. Lipid nanoparticles or AAVs comprising the oligonucleotide of the invention may be formulation as solutions for intravenous, intrathecal or intraparenchymal administration, for example.

[0421] Pharmaceutical compositions such as those of the invention may comprise naked nucleic acid sequences (e.g. oligonucleotides or vectors) or may comprise nucleic acid sequences in combination with antibodies, peptides, lipids, polymers or targeting systems. These may be delivered by any available technique. For example, the pharmaceutical composition may be introduced by needle injection, suitably intradermally, subcutaneously, intrathecally, intraparenchymally or intramuscularly. Alternatively, the pharmaceutical composition may be delivered directly across the skin using a delivery device such as particle-mediated gene delivery system. The pharmaceutical composition may be administered topically to the skin, or to mucosal surfaces for example by intranasal, oral, or intrarectal administration. The pharmaceutical composition may be administered intravenously, arterially, intradermally, intramuscularly, intraperitoneally, subcutaneously, sublingually or orally (by ingestion). The pharmaceutical composition may also appropriately be introduced by rechargeable or biodegradable polymeric devices or other devices, e.g., patches and pumps, or formulations, which provide for the extended, slow, or controlled release thereof.

[0422] In one embodiment, the pharmaceutical composition is administered intravenously.

[0423] In some embodiments, the pharmaceutical composition is formulated such that the oligonucleotide reaches a target tissue in therapeutic concentrations. Suitably the target tissue is a tissue involved in the diseases to be treated. Suitably a target tissue may be a muscle cell, cardiac cell, cardiomyocyte cell, nerve cell, brain cell, lung cell, pancreatic cell, cartilage cell (such as chondrocytes), eye cell or a bone marrow cell such as blood stem cell including for example a myeloid progenitor cell or a haematopoietic progenitor cell. In the case of the preferred diseases herein, suitably the target tissue is the muscle, heart, lung, bone marrow, cartilage, eye, pancreas or the central nervous system (CNS) including brain and spinal cord tissue.

[0424] Uptake of therapeutic oligonucleotides into a target tissue may be enhanced by several known transfection techniques, for example those including the use of transfection agents. Suitably therefore the pharmaceutical composition may comprise a transection agent. Examples of these agents include cationic agents, for example, calcium phosphate and DEAE-Dextran and lipofectants, for example, lipofectamine and transfectam.

[0425] Administration is typically in a "prophylactically effective amount" or a "therapeutically effective amount" (as the case may be, although prophylaxis may be considered therapy), this being sufficient to show benefit to the individual, e.g. an effective amount to prevent or delay onset of the disease or condition, to ameliorate one or more symptoms, to induce or prolong remission, or to delay relapse or recurrence.

[0426] The dose may be determined according to various parameters, especially according to the substance used; the age, weight and condition of the individual to be treated; the route of administration; and the required regimen. A physician will be able to determine the required route of administration and dosage for any particular individual. A typical daily dose of an active ingredient is from about 0.1 to 50 mg per kg of body weight dependent on the conditions mentioned above. The dose may be provided as a single dose or may be provided as multiple doses, for example taken at regular intervals, for example suitably, the dose may be administered daily, once every 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12, 13, 14 days, once every 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , 12 weeks, or once every 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12 months. Suitably the dose may be administered hourly. Suitably, the dose may be administered as two, three, four, five, six or more sub-doses separately at appropriate intervals throughout the day, optionally, in unit dosage forms. Typically oligonucleotides are administered at a dose of about 10-200 mg, about 10-190 mg, about 10-180 mg, about 10-170 mg, about 10-160 mg, about 10-150 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 110 mg, about 120 mg, about 130 mg, about 140 mg, about 150 mg, about 160 mg, or about 168 mg. Optionally smaller doses may be used in the range of 1 pg to 1 mg, suitably 1 pg to 10 pg of nucleic acid for particle mediated delivery and 10 pg to 1 mg for other routes.

[0427] Examples of the techniques and protocols mentioned above can be found in Remington's Pharmaceutical Sciences, 20th Edition, 2000, pub. Lippincott, Williams & Wilkins.

[0428] In some embodiments, the pharmaceutical composition comprises a further therapeutic agent, such as additional known therapeutic agents, drugs, modifications of compounds into prodrugs, and the like for alleviating, mediating, preventing, and treating the diseases, disorders, and conditions described herein. Suitably, the further therapeutic agent may include one or more active ingredients useful for the particular indication, disease, or condition being treated with the pharmaceutical composition, preferably those with activities complementary to the oligonucleotide, vector, virus particle, conjugate and / or delivery particle of the invention, where the respective activities do not adversely affect one another. Such active ingredients are suitably present in combination in amounts that are effective for the purpose intended. The further therapeutic agent may be any further agent which for example modulates the expression of the MEG3 gene, or any additional therapeutic or protective agent as described below, suitably which may be used for treating the disease that is treatable by the invention. The further therapeutic agent may be a further oligonucleotide, such as an antisense oligonucleotide. Optionally, the pharmaceutical composition may comprise two or more different oligonucleotides. Optionally, the pharmaceutical composition may further comprise one or more oligonucleotides targeting different regions of MEG3 transcripts and / or the MEG3 gene. Suitably combinations of any of the oligonucleotides described herein are envisaged.

[0429] Optionally, the one or more further oligonucleotides may be joined together and / or joined to the oligonucleotides of the invention.

[0430] Lyophilised compositions, which may be reconstituted and administered, are also within the scope of the pharmaceutical composition of the invention.

[0431] Treatments

[0432] The oligonucleotide, vector, virus particle, conjugate, delivery particle or pharmaceutical composition of the invention may be used as a medicament. Accordingly, the invention provides the oligonucleotide, vector, virus particle, conjugate, delivery particle or pharmaceutical composition of the invention for use as a medicament. Suitably therefore, the invention also provides a use of the oligonucleotide, vector, virus particle, conjugate, delivery particle or pharmaceutical composition of the invention for the manufacture of a medicament, suitably a medicament for the therapeutic applications described below.

[0433] In addition, the invention provides a method of treating or preventing a disease in a subject, wherein MEG3 is implicated in the disease, suitably a disease in which aberrant expression of the MEG3 gene is implicated, suitably a disease in which MEG3 is over-expressed or has increased expression, suitably compared to a healthy control subject, the method comprising administering to the subject an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention.

[0434] Suitably the invention provides a method of treatment of a subject having a disease, or for preventing a disease in a subject, the method comprising administering to the subject an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention. Suitably wherein the disease may be one in which MEG3 is implicated, suitably in which aberrant expression of the MEG3 gene is implicated, suitably in which MEG3 is over-expressed or has increased expression, suitably compared to a healthy control subject.

[0435] Suitably the subject is a subject in need of said treatment or prevention therapy.

[0436] The invention equally provides the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention, for use in the treatment or prevention of a disease, suitably a disease in which MEG3 is implicated, suitably in which aberrant expression of MEG3 is implicated, suitably in which MEG3 is over-expressed or has increased expression, suitably compared to a healthy control subject. Optionally for use in a method of treatment or prevention in a subject, the method optionally comprising administration of an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention to the subject.

[0437] Diseases in which MEG3 is implicated may be referred herein as target diseases and include neurodegenerative diseases, retinopathy (e.g. cancer retinopathy i.e. cancer-associated retinopathy), osteoarthritis, cardiovascular diseases, lung diseases and diabetes.

[0438] Neurodegenerative diseases treatable or preventable with the oligonucleotides of the invention include Alzheimer’s disease, spinal muscular atrophy, Parkinson's disease, Huntington’s disease, prion diseases such as Creutzfeldt-Jakob disease, motor neuron disease, amyotrophic lateral sclerosis, multiple sclerosis, frontotemporal dementia, Lewy body dementia, ataxia, spinocerebellar ataxia, multiple system atrophy, progressive supranuclear palsy, spinal muscular atrophy, spinal and bulbar muscular atrophy (SBMA) or Duchenne muscular dystrophy.

[0439] Cardiovascular diseases treatable or preventable with the invention include cardiac fibrosis.

[0440] Lung diseases treatable or preventable with the invention include obstructive pulmonary disease.

[0441] Suitably therefore, invention provides a method of treating or preventing a neurodegenerative disease, a cardiovascular disease, a lung disease, cancer retinopathy, osteoarthritis or diabetes in a subject, suitably a subject in need thereof, the method comprising administering to the subject an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention.

[0442] Suitably the invention provides a method of treatment of a subject having a disease, or for preventing a disease in a subject, the method comprising administering to the subject an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention, wherein the disease is a neurodegenerative disease, a cardiovascular disease, a lung disease, cancer retinopathy, osteoarthritis or diabetes.

[0443] The invention equally provides the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention for use in the treatment or prevention of a neurodegenerative disease, a cardiovascular disease, a lung disease, cancer retinopathy, osteoarthritis or diabetes in a subject. Optionally for use in a method of treatment or prevention in a subject, the method optionally comprising administration of an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention.

[0444] As is the case for all or substantially all neurodegenerative diseases, a defining feature of Alzheimer’s disease is neuronal cell loss through necroptosis i.e. a programmed cell death. Down-regulation of MEG3 is known to inhibit necroptosis, especially in neurons. In particular, MEG3 is known to be upregulated in subjects with Alzheimer’s diseases (Balusu et al. 2023).

[0445] Suitably therefore, the oligonucleotide, vector, virus particle, conjugate, or delivery particle of the invention may be used to treat or prevent, or be for use in the treatment or prevention of, neurodegenerative diseases by inhibiting necroptosis. Suitably, these may be used to treat or prevent, or be for use in the treatment or prevention of, Alzheimer’s disease by inhibiting necroptosis.

[0446] Suitably therefore, there is provided a method of inhibiting necroptosis in a subject in need thereof, wherein the method comprises administering to the subject an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or the pharmaceutical composition of the invention.

[0447] Suitably therefore, the invention provides a method of inhibiting necroptosis in a subject, suitably a subject in need thereof, the method comprising administering to the subject an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention. Suitably wherein a subject in need thereof is a subject with a neurodegenerative disease, suitably any neurodegenerative disease, suitably Alzheimer’s disease, spinal muscular atrophy, Parkinson's disease, Huntington’s disease, prion diseases such as Creutzfeldt-Jakob disease, motor neuron disease, amyotrophic lateral sclerosis, multiple sclerosis, frontotemporal dementia, Lewy body dementia, ataxia, spinocerebellar ataxia, multiple system atrophy, progressive supranuclear palsy, spinal muscular atrophy, spinal and bulbar muscular atrophy (SBMA) or Duchenne muscular dystrophy, most suitably Alzheimer’s disease.

[0448] Suitably the invention provides a method of inhibiting necroptosis in a subject having a disease, or for preventing a disease in a subject, the method comprising administering to the subject an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention, suitably wherein the disease is neurodegenerative disease, suitably any neurodegenerative disease, suitably Alzheimer’s disease, spinal muscular atrophy, Parkinson's disease, Huntington’s disease, prion diseases such as Creutzfeldt-Jakob disease, motor neuron disease, amyotrophic lateral sclerosis, multiple sclerosis, frontotemporal dementia, Lewy body dementia, ataxia, spinocerebellar ataxia, multiple system atrophy, progressive supranuclear palsy, spinal muscular atrophy, spinal and bulbar muscular atrophy (SBMA) or Duchenne muscular dystrophy, most suitably Alzheimer’s disease.

[0449] The invention equally provides the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention for use in the inhibition of necroptosis in a subject, the method optionally comprising administration to the subject of an effective amount of the oligonucleotide, vector, virus particle, conjugate, delivery particle, or pharmaceutical composition of the invention.

[0450] Treatment may comprise prevention, not necessarily cure such diseases. Treatment may therefore comprise a lessening of any symptoms, delay of disease progression, and / or improvement of any symptoms.

[0451] Administration of the treatment is in an "effective amount" which may be considered as a “prophylactically effective amount” or a "therapeutically effective amount" (as the case may be, although prophylaxis may be considered therapy), this being sufficient to show benefit to the individual, e.g. an effective amount to prevent or delay onset of the disease or condition, to ameliorate one or more symptoms, to induce or prolong remission, or to delay relapse or recurrence. Suitable doses for administration are provided elsewhere herein.

[0452] For example, treatment may slow the progression of disease. In some embodiments, wherein treatment is commenced in an early stage neurodegenerative disease, treatment may delay the onset of middle and / or late stage neurodegenerative disease. In some embodiments, wherein treatment is commenced during middle stage neurodegenerative disease, treatment may delay the onset of late stage neurodegenerative diseases. In some embodiments, the present disclosure pertains to: a method of ameliorating, reducing the severity of, or slowing the onset or progression of a neurodegenerative disease, a cardiovascular disease, a lung disease, cancer retinopathy, osteoarthritis or diabetes.

[0453] In one embodiment, treatment of neurodegenerative diseases with the oligonucleotide of the invention may have one or more of the following effects: improving neuronal survival; blocking, inhibiting or preventing neuronal loss; down-regulating the expression of necrosome proteins, for instance RIPK1 , RIPK3 and / or MLKL, particularly their activated forms pRIPKI , pRIPK3 and / or pMLKL; reducing inflammatory processes that cause necroptosis, particularly in the context of amyloid / tau pathology.

[0454] In one embodiment, treatment of cardiovascular diseases with the oligonucleotide of the invention may have one or more of the following effects: preventing fibrosis or diastolic dysfunction in cardiac tissue; reducing myocardial apoptosis in hypoxic condition.

[0455] In one embodiment, treatment of lung diseases with the oligonucleotide of the invention may have the following effect: preventing human pulmonary microvascular endothelial cell (HPMEC) apoptosis, e.g. in the context of chronic obstructive pulmonary disease.

[0456] In one embodiment, treatment of diabetes with the oligonucleotide of the invention may have one or more of the following effects: reducing damage to cardiac cells due to high glucose levels in blood; preventing placental cell death caused by Meg3 overexpression in the context of gestational diabetes mellitus; reducing damages to foetal endothelial function.

[0457] Suitably, treatment with a oligonucleotide of the invention reduces expression of MEG3. Suitably treatment with a oligonucleotide of the invention reduces expression of MEG3 in a subject or a target tissue in a subject by at least by 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21 %, 22%, 23%, 24%, 25%,

[0458] 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%,

[0459] 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%,

[0460] 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%,

[0461] 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%,

[0462] 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or at least 99%, compared to expression of MEG3 in a control or healthy subject or a control tissue thereof, as described in more detail elsewhere herein.

[0463] In one embodiment, the oligonucleotide of the invention reduces the level of MEG3 expression to a level that is equivalent to the expression of MEG3 in a control or healthy subject. Suitably therefore the oligonucleotide of the invention partially silences the MEG3 gene. In one embodiment, the oligonucleotide of the invention effectively silences the MEG3 gene.

[0464] Suitably the oligonucleotide is capable of reducing the level of MEG3 expression by binding to MEG3 transcripts and / or the MEG3 ger\e as described elsewhere herein. Suitably therefore, a target disease may be treated or prevented by reducing the level of MEG3 expression, suitably by RNA-directed DNA methylation (RdDM) of the MEG3 gene i.e. RITS mediated reduction or inhibition of transcription of the MEG3 gene, RNA Induced Silencing Complex (RISC) degradation of MEG3 transcripts, RNase H degradation of MEG3 transcripts or steric occlusion the RNA-lnduced Silencing Complex (RISC) degradation of MEG3 transcripts, and / or sterically occluding transcription machinery from accessing MEG3 transcripts.

[0465] In some embodiments, the present invention relates to a method of reducing the level, expression and / or activity of MEG3 transcripts, suitably in a subject. Alternatively, the present invention relates to the oligonucleotide for use in a method of reducing the level, expression and / or activity of MEG3 transcripts, suitably in a subject.

[0466] The relative level of expression of MEG3 or abundance of MEG3 transcripts may be determined by taking appropriate samples from the patient (such as samples of any target tissues mentioned elsewhere herein) treated before treatment is commenced, and once treatment has begun, and measuring the abundance of RNA by techniques such as qPCR or RNA-sequencing. Suitably therefore a determination of whether there has been a reduction in the level, expression and / or activity of MEG3 transcripts may be carried out.

[0467] The treatment may further comprise administering an additional therapeutic agent to the subject. The additional therapeutic agent may be comprised within the pharmaceutical composition as described above, or may be separate. The additional therapeutic agent may be a neuroprotective agent, an anti-inflammatory agent, an agent that regulates inflammation or immune system, an antifibrotic agent, and / or an agent that protects cells from accumulation of misfolded protein or excitatory stimuli. Suitably the additional therapeutic agent may be administered at the same time, simultaneously with, the oligonucleotide or pharmaceutical composition thereof of the invention, or at a different time. Suitably the additional therapeutic agent may act synergistically with the oligonucleotide of the invention.

[0468] The present invention relates to treatment of a disease in which MEG3 is implicated, suitably in which aberrant expression of the MEG3 gene is implicated, suitably in which MEG3 is over- expressed or has increased expression compared to a healthy control subject, such as for example a neurodegenerative disease, a cardiovascular disease, a lung disease, cancer retinopathy, osteoarthritis and diabetes, by administering an effective amount of an oligonucleotide as described herein to a subject in need thereof.

[0469] Suitably the subject has or has been diagnosed with disease in which MEG3 is implicated, suitably in which aberrant expression of the MEG3 gene is implicated, suitably in which MEG3 is over-expressed or in which expression of MEG3 is increased compared to a healthy control subject, such as for example a neurodegenerative disease, a cardiovascular disease, a lung disease, cancer retinopathy, osteoarthritis and diabetes. Suitable such disorders are listed above. In some embodiments, a subject may be diagnosed with such a disease, optionally the subject may be genetically verified to have such a disease, prior to administration of the oligonucleotide of the invention.

[0470] Suitably the subject comprises a genetic mutation or genomic alteration which leads to overexpression of MEG3, suitably which leads to the expression of MEG3 to a level which is above normal expression in a healthy subject, and which is therefore thought to be associated with disease.

[0471] In some embodiments, the present disclosure provides for a method for determining the suitability of treatment of a subject for administration of a oligonucleotide of the invention, said method comprising the steps of: i) determining the level of MEG3 expression (i.e. level of MEG3 transcripts) in a subject, ii) determining whether the level corresponds to overexpression by comparison to a suitably matched healthy control subject, and iii) optionally administering an effective amount the oligonucleotide to the subject, if the level of MEG3 expression is determined to be overexpression.

[0472] Suitably step (i) is determined by extracting a tissue sample from the subject and subjecting it to RNA quantification techniques such as qPCR, digital droplet PCR (ddPCR), or RNA- sequencing. Suitably by any known technique.

[0473] Suitably, the subject is mammalian. Suitably the subject may be an animal such as a primate (e.g., a rhesus monkey or chimpanzee), horse, dog, cat, pig, mouse, rat, rabbit, sheep, goat, ferret, or a guinea pig. Suitably the subject is human. Suitably the subject may be male or female. Suitably the subject may be of any age.

[0474] Suitably, the subject may be of any age. Suitably the subject may be between the ages of 1 month old to 100 years old, suitably between the ages of 1 years old and 90 years old, suitably between the ages of 2 years old to 80 years old, suitably between the ages of 4 years old to 75 years old.

[0475] The amount of oligonucleotide, vector, virus particle, conjugate, delivery particle of pharmaceutical composition of the invention may be administered as a single dosage form depending upon the subject treated and the particular mode of administration. Suitably, oligonucleotide, vector, virus particle, conjugate, delivery particle of pharmaceutical composition of the invention may be administered as a single dose, multiple doses or over an established period of time for example by repeated injection. Suitably, dosage regimens also may be adjusted to provide the optimum desired response (e.g., a therapeutic or prophylactic response).

[0476] Suitably the oligonucleotide, vector, virus particle, conjugate, delivery particle of pharmaceutical composition of the invention may be administered once per day, once every two days, once every three days, once per week, twice per week, once every two weeks, once per month, once every two months, once every 6 months, for example.

[0477] Exemplary single doses of the oligonucleotide may be between 0.01mg / kg and 100mg / kg of body weight. Suitable doses may therefore be: 0.05mg / kg and 90mg / kg, 0.1mg / kg and 80mg / kg, 0.5mg / kg and 70mg / kg, 1mg / kg and 60mg / kg, 2mg / kg and 50mg / kg, 5mg / kg and 40mg / kg, 10mg / kg and 30mg / kg, 12mg / kg and 20mg / kg, 13mg / kg and 17mg / kg of body weight of the oligonucleotide, or any integer values therebetween for example.

[0478] Methods of reducing MEG3 expression in a cell

[0479] Provided herein are in vitro methods of reducing the level of MEG3 expression in a cell. The cell may be any cell, suitably a cell that expresses the MEG3 gene, suitably a cell that expresses (i.e. contains) MEG3 transcripts, suitably a human cell. Suitably the cell may be any cell type mentioned here, or any other cell type.

[0480] As described elsewhere herein, reducing the level of MEG3 expression requires binding of the oligonucleotide to MEG3 transcripts and / or the MEG3 gene. The molecular mechanisms by which this occurs are also described elsewhere herein.

[0481] Suitably, there is provided a step of introducing the oligonucleotide or conjugate of the invention, or a pharmaceutical composition thereof, into a cell, under suitable conditions for the oligonucleotide to bind to MEG3 transcripts. The oligonucleotide or conjugate may be introduced into the cell by conventional means of transfection or transformation such as electroporation, lipofection, and other methods known in the art. Where the conjugate comprises the oligonucleotide linked to a delivery group such as a cell penetrating peptide (CPP), it may be sufficient to simply contact the conjugate with the cell such that the conjugate is naturally taken up by the cell by virtue of the CPP, for example by endocytosis.

[0482] Suitably, there is provided a step of introducing the virus particle or delivery particle of the invention, or a pharmaceutical composition thereof, into a cell, under suitable conditions for the oligonucleotide comprised within the virus particle or delivery particle to bind to MEG3 transcripts. Such conditions may include simply contacting the virus or delivery particle with the cell, such that the virus or delivery particle is naturally taken up by the cell, or by common methods of transfecting delivery particles such as electroporation and lipofection for example.

[0483] Suitably, there is provided a step of introducing the vector of the invention, or a pharmaceutical composition thereof, into a cell, under suitable conditions for the vector to be transcribed within the cell to produce the oligonucleotide, and for the oligonucleotide to bind to MEG3 transcripts. The vector may be introduced into the cell in the same way as oligonucleotides as described above.

[0484] Suitably, there is provided a step of introducing the virus particle, conjugate or delivery particle of the invention, or a pharmaceutical composition thereof, into a cell, under suitable conditions for the vector comprised within the virus particle, conjugate or delivery particle to be transcribed within the cell to produce the oligonucleotide, and for the oligonucleotide to bind to MEG3 transcripts.

[0485] Sutiably any of the products of the invention described herein may alternatively be introduced into the cell. Suitably the in vitro methods may be useful in research.

[0486] General techniques

[0487] The practice of the present invention will employ, unless otherwise indicated, conventional techniques of molecular biology, microbiology, cell biology, biochemistry, and immunology, which are within the skill of the art. These are described in, for example, Molecular Cloning: A Laboratory Manual, second edition (Sambrook, et al., 1989) Cold Spring Harbor Press, and Gene Transfer Vectors for Mammalian Cells (J. M. Miller and M. P. Calos, eds., 1987).

[0488] Various further aspects and embodiments of the present invention will be apparent to those skilled in the art in view of the present disclosure. Indeed, the present invention is in no way limited to the methods and materials described herein. All documents mentioned in this specification are incorporated herein by reference in their entirety.

[0489] Examples of the invention will now be described in relation to the following figures.

[0490] BRIEF DESCRIPTION OF THE DRAWINGS

[0491] Figure 1 : a cartoon illustrating the first round of screening at an oligonucleotide concentration of 250 nM of gapmer ASOs 1 to 19 against MEG3, with gapmers covering exons ENSE00002476600, ENSE00001586203 and ENSE00002440436 (A) and ASO biological activity after transfection in spinal muscular atrophy (SMA) fibroblasts (B).

[0492] Figure 2: a cartoon illustrating the second round of screening at an oligonucleotide concentration of 250 nM of gapmer ASOs against MEG3, focusing on an ‘ASO microwalk’ along exon ENSE00002440436 (A) and ASO biological activity after transfection in SMA fibroblasts (B).

[0493] Figure 3: a cell viability assay performed with a panel of ASOs targeting the three exons of MEG3-226 in SMA fibroblasts. Cells were transfected at a concentration of 1 pM and showed no sign of ASO cytotoxicity after 24h treatment.

[0494] Figure 4: biological activity of ASOs 1-19 after transfection at 250 nM in AC-16 cardiac cells.

[0495] Figure 5: dose-response curves obtained 36h post-transfection for selected ASOs 8, 15, 16, and 18 in SMA fibroblasts, which show potent knockdown already at 1 nM concentration.

[0496] Figure 6: a dose-dependent, siRNA-mediated knockdown of MEG3 in SMA fibroblasts, with three increasing concentrations of the siRNA “ASO 18” and a treatment time of 36h or 72h.

[0497] EXAMPLES

[0498] The MEG3 transcript sequence was retrieved from the Ensembl database, and used to design oligonucleotide sequences. Accordingly, Ensembl nomenclature is being used to identify specific transcripts and exons. The skilled person will be aware that NCBI nomenclature could also be used, in which case transcripts would have a different IDs starting with “NR_”. Materials and Methods

[0499] Human LncRNA MEG3 sequence was identified from Ensembl (Release 110). The inventors designed antisense gapmer oligonucleotides, short “ASOs”, against MEG-226 transcript (ID ENST00000556407.5) and its exons ENSE00002476600, ENSE00001586203 and

[0500] ENSE00002440436. All ASOs were 20 nucleotides length with “5-10-5” scaffold, i.e. , a central 10-nt DNA core and 5-nt wings modified with 2’-O-methoxyethyl (2’MOE) modified RNA. Oligonucleotide backbones were fully phosphorothioated. All cytosines, i.e. 2’MOE RNA and DNA, are 5-methyl substituted; in other words, all cytosine nucleobases are 5-methylcytosines (5MeC). All sequences are provided herein as 5’ to 3’.

[0501] Oligonucleotide synthesis was conducted on a H-16 synthesizer (K&A Labs Gmbh) with the phosphoramidite chemistry. Pre-packed 1 pmol Universal Support Columns were used (LGC, 1000 Angstrom pore-size, cat. MLX1-3500-1). Cycle sequence was: Detritylation, Coupling, Sulfurization, and Capping. Detritylation was carried out with a solution of 3% trichloroacetic acid in dichloromethane (Merck, cat. L020000). 2 -O-MOE (i.e. 2’MOE) phosphoramidites and DNA phosphoramidites were purchases from ChemGenes and prepared at 0.1 M concentration in dry acetonitrile. A 0.3 M solution of 5-(Benzylthio)-1 H-tetrazole in dry acetonitrile (ChemGenes, cat. RN-1452) was used to activate the phosphoramidites for coupling. Sulfurization was carried out with a 0.05 M solution of 3-((N,N- dimethylaminomethylidene)amino)-3H-1 , 2, 4-dithiazole-5-thione (ChemGenes, cat. RN- 1588) in a 1 :1 mixture of pyridine / acetonitrile. Capping of failed sequences was achieved with a 1 :1 mixture of Capping A (10% acetic anhydride, 10 % 2, 6-lutidine, and 80% tetrahydrofuran, ChemGenes cat. RN-1458) and Capping B (16% N-methylimidazole in tetrahydrofuran, ChemGenes cat. RN-7776). A final detritylation was included post-synthesis to generate 5’- hydroxy oligonucleotides. and characterisation

[0502] Solid supports were transferred into screw-cap tubes and suspended in a 1 ml solution of 25% aqueous ammonia (Merck). Oligonucleotide cleavage and deprotection was carried out by heating the suspension at 65°C for 8 hours. Tubes were allowed to cool down to room temperature, spun down, and placed in a rotary vacuum concentrator. Crude oligonucleotides were concentrated to dryness. Oligonucleotides were redissolved in 400 l RNase-free water and separated from solid residues by centrifugation. Supernatants were transferred into fresh screw-cap tubes, made 0.3 M sodium acetate using a stock of 3 M sodium acetate, and mixed with 4 volumes of molecular biology grade ethanol. Suspensions were cooled overnight at -20°C and then centrifuged at 4°C for 15 mn at 12500 g. Supernatants were discarded, pellets were briefly dried on benchtop for 15 mn, washed with 400 pl of ice-chilled 90% ethanol, and immediately re-centrifuged for 10 mn at 12500 g and 4°C temperature. Supernatants were discarded, pellets were briefly dried on benchtop for 15 mn and in a Speedvac for 1 mn. Pellets were then redissolved in 400 pl of RNAse free water with gentle shaking on a thermoshaker for over 30 min.

[0503] Oligonucleotide integrity was confirmed by liquid chromatography-mass spectrometry analysis on a Waters SQD2 system coupled to a Waters Acquity HPLC fitted with a C-18 column (Agilent AdvanceBio, 2.1 x 50 mm, 2.7 pm). LC-MS runs were carried out at 0.5 ml / min, column temperature was 60°C, and mobile phase was made of Buffer A, 0.4 M hexafluoroisopropanol, 15 mM triethylamine in water, and buffer B, methanol. Mass range m / z was 750-3000.

[0504] Cell culture experiments

[0505] Spinal muscular atrophy (SMA) donor fibroblasts (Coriell Institute cat. GM03813) were grown in DMEM-Glutamax supplemented with 10 % fetal bovine serum and antibiotic-antimycotic solution. Cells were kept in culture at 37°C, 5% CO2. For experiments, cells were plated at 50’000 cells / well in 1 ml medium in a 24-well plate. Cells were allowed to re-attach overnight before transfection with Lipofectamine 2000. The total volume of oligonucleotide treatment solutions was 100 pl per well. Oligonucleotides were pre-diluted in 50 pl OptiMEM medium immediately prior to mixing with 49 pl OptiMEM containing 1 pl Lipofectamine 2000. Solutions were incubated for 20 min and added into cell culture wells. Treatments were allowed to run for 36 hours.

[0506] AC- 16 cells (Merck) were grown in DMEM-Glutamax supplemented with 10 % fetal bovine serum and antibiotic-antimycotic solution. Cells were kept in culture at 37°C, 5% CO2. For experiments, cells were plated at 35’000 cells / well in 0.5 ml medium in a 24-well plate. Cells were allowed to re-attach overnight before transfection with Lipofectamine 2000. The total volume of oligonucleotide treatment solutions was 100 pl per well. Oligonucleotides were prediluted in 50 pl OptiMEM medium immediately prior to mixing with 49 pl OptiMEM containing 1 l Lipofectamine 2000. Solutions were incubated for 20 min and added into cell culture wells. Treatments were allowed to run for 36 hours.

[0507] Total RNA extraction and reverse transcription

[0508] Cells were washed one time with 500 pl phosphate-buffered saline (PBS) solution. RNA extraction was carried out using Qiagen RNEasy kit according to manufacturer’s instructions and including the optional DNase I treatment step. cDNA was generated using PrimeScript RT kit (Takara, cat. RR037A) with 300-500 ng RNA as input, or with the High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems #4368814) with 400 ng RNA as input. cDNA samples were diluted 1 / 5 or 1 / 8 v / v in RNase-free water prior to use in real time PCR. Real time PCR

[0509] RT-qPCR reactions were prepared with the FAST SYBR Mix (Applied Biosystems, cat. 4385612) or the SYBR Green™ Gene Expression Master Mix (Applied Biosystems # 4369016) and run and analysed on a StepOnePlus™ real-time PCR system (Applied Biosystems) according to manufacturer’s instructions. Gene-specific primers for transcript amplification were synthesized by Integrated DNA Technologies and are listed in the table below. B2M or TBP were used as house-keeping control. Cell

[0510] Cell viability assays were conducted in technical triplicates in 96 well-plate format, using Promega’s RealTime-Glo MT Cell Viability Assay (Promega G9712) and according to manufacturer’s instructions. Cells were plated at 10 000 cells / well in 200 pl medium in a 96- well plate and oligonucleotides delivered by Lipofectamine 2000 as in activity studies, at a higher concentration of 1 uM concentration. Viability was measured 24 h post treatment in a microplate reader.

[0511] Dosecurves

[0512] SMA fibroblast cells (GM03813) were plated at 20 000 cells / well in 400 ul of medium in a CytoOne 48-well plate. Remaining method was the same described under “Cell culture experiments”. ASOs 8, 15, 16 and 18 were selected for the dose-response experiments and transfected at 1 , 10, 25, 50, 125, 250, 500, 750, and 1000 nM over a 36-hour period. siRNA transfection experiment

[0513] For siRNA experiments, an siRNA molecule based on ASO 18 was purchased from Integrated DNA Technologies. The antisense strand had the same nucleotidic sequence as ASO18, but was a full RNA phosphodiester molecule and had a 3’ dTdT DNA overhang. The sense strand was the reverse complement of ASO 18 and was also full RNA phosphodiester with a 3’ dTdT DNA overhang.

[0514] Antisense sequence (5’-3’): rUrUrArGrGrUrArArGrArGrGrGrArCrArGrCrllrGdTdT (SEQ ID NO: 127); sense sequence (5’-3’): rCrArGrCrUrGrUrCrCrCrUrCrUrUrArCrCrllrArAdTdT

[0515] (SEQ ID NO: 128); with rA, rC, rG, or rU indicating a natural RNA ribonucleotide; and dT indicating a deoxythymidine.

[0516] SMA fibroblast cells (GM03813) were plated at 50 000 cells / well in 1 ml of medium in a 24- well plate. siRNA transfection occurred 24h after plating using RNAiMax (Invitrogen 13778- 075) as per the manufacturer’s instructions. siRNA was diluted in 50 pl medium prior to mixing with 47 pl Opti-MEM containing 3 pl of RNAiMAX reagent. Remaining method was the same described under “Cell culture experiments”. siRNA treatments were performed at 1 , 10 and 40 nM concentration and allowed to run for 36h or 72h. Results

[0517] Design of qapmer ASO sequences

[0518] A great number of MEG3 splicing variants are reported in genomic databases. Ensembl lists a total of 50 variants, while NCBI reports 15 variants. The design of nucleic acid sequences targeting simultaneously many MEG3 splicing variants is challenging, as many exons are specific for single or small sets of variants. For instance, an shRNA targeting MEG3 widely used in literature (e.g., in Balusu et al. 2023, Zhang 2019, and Bi et al. 2020) only targets a few splicing variants and does not bind to Ensembl’s MEG3 reference (canonical) transcript, MEG-226. MEG-226 is composed of three exons ENSE00002476600, ENSE00001586203 and

[0519] ENSE00002440436. While ASOs targeting ENSE00002476600 would be specific for MEG- 226, ASOs binding to ENSE00001586203 and ENSE00002440436 can theoretically bind an extended number of splicing variants. The inventors hypothesised that such ASOs would lead to improved MEG3 knock-down. Specifically, ENSE00001586203 is found in the following MEG3 transcript variants:

[0520] The sequence of ENSE00001586203 is also found in full as part of a bigger exon in MEG3 splicing variants ENST00000783980.1 (MEG3-264) and ENST00000783985.1 (MEG3-269). A part of the sequence of ENSE00001586203 is also found in MEG3 splicing variant ENST00000648138.1 (MEG3-230).

[0521] ENSE00002440436 is found in the following MEG3 transcript variants:

[0522] A part of the sequence of ENSE00002440436 is also found in MEG3 splicing variant ENST00000649036.2 (MEG3-238).

[0523] Screening of exons ENSE00002476600, ENSE00001586203 and ENSE00002440436 The inventors first designed and synthesised ASOs 1-19 (Figure 1A) for targeting of exons ENSE00002476600 (ASOs 1-7), ENSE00001586203 (ASOs 8-13) and ENSE00002440436 (ASOs 14-19). All ASOs were tested in SMA Type II fibroblasts (Coriell GM03813), a cell line suitable for oligonucleotide transfection and for which the inventors have verified Meg3 expression. ASOs were transfected at a concentration of 250 nM and knock-down of Meg3 was assessed by RT-qPCR. Three negative controls, i.e. randomised or scrambled ASO sequences, were also included to verify assay specificity.

[0524] Different potencies and patterns were obtained for ASOs 1-19 (Figure 1B). ASOs 1 to 7 targeted ENSE00002476600, with ASO5 being particularly effective, achieving 65% knockdown. ASOs 8 to 13 targeting short exon ENSE00001586203 were even more potent, with knock-down values of up to 84% (ASO13). Targeting of the 3’ end of exon ENSE00001586203 was particularly effective with ASO12 and ASO13 achieving 78% and 84% knock-down, respectively. Finally, ASOs 14 to 19 targeted ENSE00002440436 and had the most activity, with knockdown values of up to 96% (ASO16). It is notable that substantially all ASOs targeting this exon were highly potent, with all but the most 3’ ASO, ASO19, achieving between 72% and 96% knock-down. ASOs 15, 16 and 18 in particular were remarkably potent, with knockdown values between 90% and 96%.

[0525] Microwalk on exon ENSE00002440436

[0526] Based on the results obtained with ASOs 14-19, we designed and synthesised additional ASO sequences 20-31 for enhanced coverage of ENSE00002440426 (Figure 2A). These ASOs were tested alongside ASOs 14-19 in the same cell assay (Figure 2B). We found all new ASOs to be active. Remarkably, ASOs 20, 21 , 22, 23, 26, 27, 28, 30, and 31 all resulted in more than 90% knockdown. ASOs 24, 25 were slightly less potent with 81 and 73% knockdown, respectively, while ASO 29 was the least active of the set with 66% knockdown.

[0527] Further testing of oligonucleotides and cytotoxicity screening

[0528] The inventors next tested a sample of ASOs at a higher concentration of 1 pm, four-fold higher than that used in experiments mentioned so far to assess the cytotoxicity of the ASOs (Figure 3). Results showed no cytotoxicity for any of the tested ASOs.

[0529] Dose-response curves

[0530] ASOs 8, 15, 16, 18 all showed a potent knockdown and were investigated further in doseresponse experiments. ASO 8 and 15 showed approximately 40% reduction in MEG3 levels at the first dose tested of 1 nM, and a steep reduction in MEG3 levels between 10 and 50 nM with a plateau reached at 250 nM (Figure 5). ASO 16 and ASO 18 were the most active of the set, with a 60 % reduction in Meg3 levels attained at 1 nM concentration, and knock-down values rising to 80% (ASO16) or 90% (ASO18) at 250 nM concentration and beyond. siRNA experiments

[0531] An siRNA form of ASO18 was transfected into SMA fibroblast cells. The oligonucleotide led to approximately 70-90% reduction in MEG3 expression over a 72 hour period at the concentrations tested (Figure 6).

[0532] Testing of oligonucleotides in AC16 cardiomyocyte cells

[0533] Next, MEG3 knock-down was performed in cardiac cells, specifically cardiomyocyte cells (Figure 4). Trends in the levels of knock-down were similar to those attained in SMA cells.

[0534] Conclusion

[0535] It is believed that the efficacy demonstrated by ASOs in the Examples herein may egually be achieved by other oligonucleotides such as siRNAs or shRNAs, for examples, as well as any other types of oligonucleotides, particularly any other types of oligonucleotides that are commonly used therapeutic modalities.

[0536] REFERENCES

[0537] Balusu, Sriram, et al. "MEG3 activates necroptosis in human neuron xenografts modeling Alzheimer’s disease." Science 381.6663 (2023): 1176-1182.

[0538] Bi, Hui, et al. "Long noncoding RNA (IncRNA) maternally expressed gene 3 (MEG3) participates in chronic obstructive pulmonary disease through regulating human pulmonary microvascular endothelial cell apoptosis." Medical science monitor: international medical journal of experimental and clinical research 26 (2020): e920793-1.

[0539] Piccoli, Maria-Teresa, et al. "Inhibition of the cardiac fibroblast-enriched IncRNA Meg3 prevents cardiac fibrosis and diastolic dysfunction." Circulation research 121.5 (2017): 575- 583.

[0540] Zhang, Hailing. "Mechanism associated with aberrant IncRNA MEG3 expression in gestational diabetes mellitus." Experimental and Therapeutic Medicine 18.5 (2019): 3699-3706.

[0541] SEQUENCES

[0542] The first column in the tables below contain SEQ ID NOs. Target regions Oligonucleotide sequences (unmodified)

[0543] Primers

[0544] Target regions (continued)

[0545] ASO sequences

[0546] *Each of SEQ ID NO: 57-87 below contains a central 10-nt DNA core (underlined) surrounded by 5-nt RNA wings, wherein the wings consist of 2’-O-methoxyethyl (2’MOE) modified RNA nucleotides. Oligonucleotide backbones were fully phosphorothioated; in other words, every 19 internucleoside linkage is a phosphorothioate (PS) internucleoside linkage. All cytosines, i.e. 2’MOE RNA and DNA, are 5-methyl substituted; in other words, all cytosine nucleobases are 5-methylcytosines (5MeC). All sequences are provided herein as 5’ to 3’.

[0547] Target sequences

[0548] Target regions (continued) siRNA sequences

[0549] In the sequences below, “r” before a nucleotide represents that the nucleotide after “r” is a ribonucleotide, e.g. rA is a ribonucleotide adenine. Nucleotides without “r” before it are deoxyribonucleotides. siRNA sequences

[0550] In the sequences below, each nucleotide is a ribonucleotide.

Claims

1. CLAIMS1. An oligonucleotide for reducing the level of MEG3 expression in a cell, wherein the oligonucleotide is capable of binding to a target sequence at least partially within a region of a MEG3 transcript selected from the group consisting of: SEQ ID NO: 3, SEQ ID NO: 2 and SEQ ID NO: 1.

2. The oligonucleotide of claim 1 , wherein the region of the MEG3 transcript is selected from the group consisting of: SEQ ID NO: 2 and SEQ ID NO: 3, preferably wherein the region of the MEG3 transcript is selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 3.

3. The oligonucleotide of any of claims 1 or 2, wherein the region of the MEG3 transcript is SEQ ID NO: 3, or wherein the region of the MEG3 transcript is selected from the group consisting of: SEQ ID NO: 4 and SEQ ID NO: 5.

4. The oligonucleotide of any preceding claim, wherein the region of the MEG3 transcript is selected from the group consisting of: SEQ ID NO: 5, preferably wherein the region of the MEG3 transcript is selected from the group consisting of: SEQ ID NO: 6, more preferably wherein the region of the MEG3 transcript is selected from the group consisting of: SEQ ID NO: 7, 8 and 9.

5. The oligonucleotide of any preceding claim, wherein the oligonucleotide comprises a region of complementarity to the target sequence, wherein the complementarity may be partial or complete complementarity.

6. The oligonucleotide of claim 5, wherein the region of complementarity is at least 5 nucleotides in length, preferably wherein the region of complementarity is no more than 120 nucleotides in length.

7. The oligonucleotide of any of claims 5 or 6, wherein the region of complementarity is at least 50% complementary to the target sequence.

8. The oligonucleotide of any preceding claim, wherein the oligonucleotide comprises ribonucleotides and / or deoxyribonucleotides.

9. The oligonucleotide of any preceding claim, wherein the oligonucleotide is singlestranded, double-stranded or comprises both single-stranded and double-stranded regions.

10. The oligonucleotide of any preceding claim, wherein the oligonucleotide is doublestranded and consists of ribonucleotides, preferably wherein the oligonucleotide is a short interfering RNA (siRNA).

11. The oligonucleotide of any of claims 1-9, wherein the oligonucleotide is single-stranded, preferably wherein the oligonucleotide is a short hairpin RNA (shRNA), or an antisense oligonucleotide (ASO), more preferably wherein the oligonucleotide is a mixmer ASO or a gapmer ASO.

12. The oligonucleotide of any preceding claim, wherein the oligonucleotide is capable of promoting degradation of MEG3 transcripts.

13. The oligonucleotide of claim 12, wherein the degradation is mediated by RNase H.

14. The oligonucleotide of any preceding claim, wherein the oligonucleotide comprises one or more modified nucleotides, preferably one or more 2’-modified nucleotides, one or more 2-modified nucleotides, one or more 3-modified nucleotides, one or more 4- modified nucleotides, one or more 5-modified nucleotides, one or more 6-modified nucleotides, one or more 7-modified nucleotides and / or one or more 8-modified nucleotides, more preferably selected from the group consisting of: 2’-O-methyl (2’OMe), 2’-O-methoxyethyl (2’MOE), 2’-fluoro (2’F), 2’-O-aminopropyl (2’OAP), 2’-O- dimethylaminoethyl (2’ODMAE), 2’-O-dimethylaminopropyl (2’ODMAP), 2’-O- dimethylaminoethyloxyethyl (2’ODMAEOE), 2’-O-N-methylacetamido (2’ONMA), 2'- dimethylaminooxyethoxy (2’-DMAO), 2’-dimethylaminoethoxyethoxy (2’-DMAEOE), 2’- amino, 2-amino (such as 2-amino pyrimidines such as 2-aminopyrimidine, 2- aminothymine, 2-aminocytosine and 2-aminouracil), 2-hydroxyl (such as 2-hydroxyl purines such as 2-hydroxypurine, 2-hydroxyadenine and 2-hydroxyguanine), 2-propyl , 2- thio, 2-fluoro (such as 2-fluoro pyrimidines such as 2-fluoropyrimidine, 2-fluorothymine, 2-fluorocytosine and 2-fluorouracil), 3-deaza, 4-thio (such as 4-thio pyrimidines such as4-thiopyrimidine, 4-thiothymine, 4-th iocytosine and 4-thiouracil), 5-methyl (such as 5- methylcytosine, i.e. 5MeC), 5-halo, 5-propynyl, 5-hydroxymethyl, 5-bromo, 5- trifluoromethyl, 6-methyl, 6-azo, 7-methyl, 7-deaza, 8-halo, 8-amino, 8-thiol, 8-thioalkyl and 8-hydroxyl nucleotides, most preferably wherein the mixmer ASO or gapmer ASO comprises one or more 2’-O-methoxyethyl (2’MOE) and one or more 5-methyl (such as5-methylcytosine (5MeC)) nucleotides.

15. The oligonucleotide of any preceding claim, wherein the oligonucleotide comprises one or more 2’-4’ bicyclic modified nucleotides, preferably selected from the group consisting of: locked nucleotides (locked nucleic acid monomers i.e. LNA monomers), 2’, 4’- constrained 2’-O-methoxyethyl (c2’MOE) nucleotides, 2’,4’-constrained 2’-0-ethyl (cEt) nucleotides, and 2’-O,4’-C-ethylene-bridged (ENA) nucleotides (bridged nucleic acids), tricyclo nucleotides.

16. The oligonucleotide of any preceding claim, wherein the oligonucleotide comprises one or more modified internucleoside linkages, preferably selected from the group consisting of: a phosphotriester linkage, a phosphorothioate (PS) internucleoside linkage, a methylphosphonate internucleoside linkage, a phosphorodithioate internucleoside linkage, a boranophosphate internucleoside linkage, a phosphoramidate internucleoside linkage, a phosphorodiamidate internucleoside linkage, a thiophosphoramidate linkage, a thiophosphorodiamidate linkage, a phosphonocarboxylate internucleoside linkage, a phosphonoacetate internucleoside linkage, an aminoalkylphosphotriester internucleoside linkage, a methyl or other alkyl phosphonate internucleoside linkage, (such as a 3’- alkylene phosphonate internucleoside linkage or a 5’-alkylene phosphonate internucleoside linkage), a phosphinate internucleoside linkage, a phosphoramidate internucleoside linkage (such as a 3’-amino phosphoramidate internucleoside linkage or an aminoalkylphosphoramidate internucleoside linkage), a thionophosphoramidate internucleoside linkage, a thionoalkylphosphotriester internucleoside linkage, a selenophosphate internucleoside linkage, preferably wherein the mixmer ASO or gapmer ASO comprises one or more phosphorothioate (PS) internucleoside internucleoside linkages.

17. The oligonucleotide of any preceding claim, wherein the oligonucleotide comprises one or more artificial nucleotides, preferably selected from the group consisting of: phosphorodiamidate morpholino monomers and peptide nucleic acid monomers.

18. The oligonucleotide of any preceding claim, wherein the oligonucleotide comprises one or more 2’-O-methoxyethyl (2’MOE) modified nucleotides, one or more 5-methylcytosine (5MeC) nucleotides, and one or more phosphorothioate (PS) internucleoside linkages.

19. The oligonucleotide of any of claims 1-9 and 11-18, wherein the oligonucleotide is a gapmer ASO and preferably comprises the structure 5’-A-B-C-3’, wherein:A consists of ribonucleotides or deoxyribonucleotides,B consists of deoxyribonucleotides,C consists of ribonucleotides or deoxyribonucleotides; preferably whereinA consists of ribonucleotidesB consists of deoxyribonucleotides C consists of ribonucleotides.

20. The oligonucleotide of claim 19, wherein:A comprises 5 nucleotides,B comprises 10 nucleotides, and C comprises 5 nucleotides.

21. The oligonucleotide of any of claims 1-20, wherein the oligonucleotide comprises a sequence with 70% or greater identity to a sequence selected from the group consisting of: SEQ ID NO: 10-40, or a functional variant thereof.

22. The oligonucleotide of any of claims 1-21, wherein the oligonucleotide consists of a sequence selected from the group consisting of: SEQ ID NO: 57-87.

23. A vector comprising a nucleic acid encoding the oligonucleotide of any of claims 1-22.

24. A virus particle comprising the oligonucleotide of any of claims 1-22 or the vector of claim 23.

25. A conjugate comprising the oligonucleotide any of claims 1-22 covalently linked to a delivery group, preferably wherein the delivery group is a cell penetrating peptide.

26. A delivery particle comprising the oligonucleotide of any of claims 1-22, the vector of claim 23, or the conjugate of claim 25, preferably wherein the delivery particle is a nanoparticle or a vesicle, more preferably a lipid nanoparticle (LNP).

27. A pharmaceutical composition comprising the oligonucleotide of any of claims 1-22, vector of claim 23, virus particle of claim 24, conjugate of claim 25, or delivery particle of claim 26, and one or more pharmaceutically acceptable excipients.

28. The oligonucleotide of any of claims 1-22, vector of claim 23, virus particle of claim 24, conjugate of claim 25, delivery particle of claim 26, or pharmaceutical composition of claim 27, for use as a medicament.

29. The oligonucleotide of any of claims 1-22, vector of claim 23, virus particle of claim 24, conjugate of claim 25, delivery particle of claim 26, or pharmaceutical composition of claim 27, for use in the treatment or prevention of a neurodegenerative disease, preferably Alzheimer’s disease, spinal muscular atrophy, Parkinson's disease, Huntington’s disease, prion diseases such as Creutzfeldt-Jakob disease, motor neuron disease, amyotrophic lateral sclerosis, multiple sclerosis, frontotemporal dementia, Lewy body dementia, ataxia, spinocerebellar ataxia, multiple system atrophy, progressive supranuclear palsy, spinal muscular atrophy, spinal and bulbar muscular atrophy (SBMA) or Duchenne muscular dystrophy.

30. The oligonucleotide of any of claims 1-22, vector of claim 23, virus particle of claim 24, conjugate of claim 25, delivery particle of claim 26, or pharmaceutical composition of claim 27 for use according to claim 29, wherein the neurodegenerative disease is treated or prevented by the inhibition of necroptosis.

31. The oligonucleotide of any of claims 1-22, vector of claim 23, virus particle of claim 24, conjugate of claim 25, delivery particle of claim 26, or pharmaceutical composition of claim 27, for use in the treatment or prevention of a cardiovascular disease, preferably cardiac fibrosis.

32. The oligonucleotide of any of claims 1-22, vector of claim 23, virus particle of claim 24, conjugate of claim 25, delivery particle of claim 26, or pharmaceutical composition of claim 27, for use in the treatment or prevention of a lung disease, preferably obstructive pulmonary disease.

33. The oligonucleotide of any of claims 1-22, vector of claim 23, virus particle of claim 24, conjugate of claim 25, delivery particle of claim 26, or pharmaceutical composition of claim 27, for use in the treatment or prevention of diabetes.

34. The oligonucleotide of any of claims 1-22, vector of claim 23, virus particle of claim 24, conjugate of claim 25, delivery particle of claim 26, or pharmaceutical composition of claim 27, for use according to any of claims 29-33, wherein the disease is treated or prevented by reducing the level of MEG3 expression, preferably by RNase H or the RNA-lnduced Silencing Complex (RISC) degradation of MEG3 transcripts.

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