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BR-112021009213-B1 - Nucleic acids for inhibition of LPA expression in a cell and composition comprising the same.

BR112021009213B1BR 112021009213 B1BR112021009213 B1BR 112021009213B1BR-112021009213-B1

Abstract

NUCLEIC ACIDS FOR INHIBITING LPA EXPRESSION IN A CELL. The present invention relates to products and compositions and their uses. In particular, the invention relates to nucleic acid products that interfere with or inhibit the expression of the LPA gene, preferably for use as treatment, prevention or reduction of the risk of suffering from cardiovascular diseases, such as coronary artery disease or aortic stenosis or stroke or any other disorder, pathology or syndrome linked to elevated levels of Lp(a)-containing particles.

Inventors

  • DAVID ANTHONY RIDER
  • Lucas Bethge
  • Christian Frauendorf
  • ADRIEN WEINGAERTNER
  • Judith HAUPTMANN
  • Sibylle DAMES
  • Steffen Schubert
  • STEPHAN TENBAUM

Assignees

  • SILENCE THERAPEUTICS GMBH

Dates

Publication Date
20260317
Application Date
20191113
Priority Date
20190514

Claims (13)

  1. 1. Nucleic acid for inhibiting LPA expression in a cell, characterized by comprising a first strand and a second strand, wherein the first strand comprises a nucleotide sequence of 5' mA (ps) fU (ps) mA fA mC fU mC fU mG fU mC fC mA fU mU fA mC (ps) fC (ps) mG 3' (SEQ ID NO: 165), and wherein the second strand comprises a nucleotide sequence of 5' mC mG mG mU mA mA fU fG fG mA mC mA mG mA mG mU mU (ps) mA (ps) mU 3' (SEQ ID NO: 163); wherein fA, fC, fG and fU denote 2'-deoxy-2'-fluororibonucleotides; mA, mC, mG and mU denote 2'-O-methyl ribonucleotides; and (ps) denotes a phosphorothioate linkage.
  2. 2. Nucleic acid, according to claim 1, characterized in that the first strand consists of a nucleotide sequence of 5' mA (ps) fU (ps) mA fA mC fU mC fU mG fU mC fC mA fU mU fA mC (ps) fC (ps) mG 3' (SEQ ID NO: 165); wherein fA, fC, fG and fU denote 2'-deoxy-2'-fluororibonucleotides; mA, mC, mG and mU denote 2'-O-methyl ribonucleotides; and (ps) denotes a phosphorothioate linkage.
  3. 3. Nucleic acid, according to claim 1 or 2, characterized in that the second strand consists of a nucleotide sequence of 5' mC mG mG mU mA mA fU fG fG mA mC mA mG mA mG mU mU (ps) mA (ps) mU 3' (SEQ ID NO: 163); wherein fA, fC, fG and fU denote 2'-deoxy-2'-fluororibonucleotides; mA, mC, mG and mU denote 2'-O-methyl ribonucleotides; and (ps) denotes a phosphorothioate linkage.
  4. 4. Nucleic acid, according to any one of claims 1 to 3, characterized in that the nucleic acid is conjugated to a ligand.
  5. 5. Conjugated nucleic acid, according to claim 4, characterized in that the ligand comprises (i) one or more N-acetylgalactosamine (GalNAc) moieties, and (ii) a connector, wherein the connector conjugates GalNAc to the nucleic acid.
  6. 6. Conjugated nucleic acid, according to claim 4, characterized in that the ligand comprises a compound of formula (I):[S-X1-P-X2]3-A-X3- (I)wherein:S represents a saccharide, wherein the saccharide is N-acetylgalactosamine;X1 represents C3-C6 alkylene or (-CH2-CH2-O)m(-CH2)2- wherein m is 1, 2, or 3;P is a phosphate or modified phosphate, preferably a thiophosphate;X2 is alkylene or an alkylene ether of formula (-CH2)n-O-CH2-, wherein n = 1-6;A is a branching unit;X3 represents a bridging unit;wherein the nucleic acid is conjugated to X3 by means of a phosphate or modified phosphate.
  7. 7. Conjugated nucleic acid, according to claim 5, characterized in that the modified phosphate is a thiophosphate.
  8. 8. Conjugated nucleic acid, according to claim 6, characterized in that the nucleic acid is conjugated to X3 via the 5' end of the second strand.
  9. 9. Conjugated nucleic acid, according to claim 4, characterized in that the conjugated nucleic acid has the following structure: where Z is the nucleic acid.
  10. 10. Conjugated nucleic acid, according to claim 9, characterized in that the nucleic acid is conjugated to the ligand via the 5' end of the second strand.
  11. 11. Conjugated nucleic acid, according to claim 4, characterized in that the second strand comprises a nucleotide sequence of 5' [ST23 (ps)]3 C6XLT (ps) mC mG mG mU mA mA fU fG fG mA mC mA mG mA mG mU mU (ps) mA (ps) mU 3' (SEQ ID NO: 164); wherein fA, fC, fG and fU denote 2'-deoxy-2'-fluoro ribonucleotides; mA, mC, mG and mU denote 2'-O-methyl ribonucleotides; (ps) denotes a phosphorothioate linkage; and [ST23 (ps)]3 C6XLT (ps) denotes
  12. 12. Conjugated nucleic acid, according to claim 11, characterized in that the second strand consists of a nucleotide sequence of 5' [ST23 (ps)]3 C6XLT (ps) mC mG mG mU mA mA fU fG fG mA mC mA mG mA mG mU mU (ps) mA (ps) mU 3' (SEQ ID NO: 164); wherein fA, fC, fG and fU denote 2'-deoxy-2'-fluororibonucleotides; mA, mC, mG and mU denote 2'-O-methyl ribonucleotides; (ps) denotes a phosphorothioate linkage; and [ST23 (ps)]3 C6XLT (ps) denotes
  13. 13. Composition, characterized by comprising a nucleic acid or a conjugated nucleic acid as defined in any of claims 1 to 12 and a vehicle and/or a diluent and/or a buffer and/or a preservative.

Description

FIELD OF THE INVENTION [001] The present invention relates to products and compositions and their uses. In particular, the invention relates to nucleic acid products that interfere with or inhibit the expression of the LPA gene. This therapeutic Lp(a) reduction therapy serves to prevent and reduce the risk of stroke, atherosclerosis, thrombosis and cardiovascular diseases, such as coronary heart disease and aortic stenosis, or any other disorder, pathology or syndrome linked to elevated levels of Lp(a)-containing particles. FUNDAMENTALS OF INVENTION [002] Double-stranded RNA (dsRNA) capable of complementarily detaching expressed mRNA has been shown to be able to block gene expression (Fire et al., 1998, Nature. February 19, 1998; 391(6669):806-11 and Elbashir et al., 2001, Nature. May 24, 2001; 411(6836):494-8) through a mechanism termed RNA interference (RNAi). Short dsRNAs direct gene-specific post-transcriptional silencing in many organisms, including vertebrates, and have become a useful tool for studying gene function. RNAi is mediated by the RNA-induced silencing complex (RISC), a sequence-specific multicomponent nuclease that degrades messenger RNAs homologous to the silencing trigger carried in the RISC complex. Interfering RNA (referred to as iRNA in this document), such as siRNAs, antisense RNA, and microRNAs, are oligonucleotides that prevent protein formation by silencing genes; that is, they inhibit the translation of protein genes through the degradation of mRNA molecules. Gene silencing agents are becoming increasingly important for therapeutic applications in medicine. [003] According to Watts and Corey in the Journal of Pathology (2012; Vol 226, p 365-379), there are algorithms that can be used to design nucleic acid silencing triggers, but they all have serious limitations. Several experimental methods may be needed to identify potent iRNAs, as the algorithms do not take into account factors such as the tertiary structure of the target mRNA or the involvement of RNA-binding proteins. Therefore, discovering a potent nucleic acid silencing trigger with minimal off-target effects is a complex process. For the pharmaceutical development of these highly charged molecules, it is necessary that they can be synthesized economically, distributed to target tissues, enter cells, and function within acceptable toxicity limits. [004] Lp(a) particles are heterogeneous low-density lipoprotein particles predominantly expressed in the liver (Witztum and Ginsberg, J. Lipid Res. 2016 Mar; 57(3):336-9). They are composed of Apolipoprotein(a) (Apo(a) or Lp(a) encoded by the LPA gene) linked to an LDL-like particle via the ApoB polypeptide. Genetically defined elevated serum levels of Lp(a) are unaffected by diet and exercise and are associated with a higher risk of cardiovascular disease through the associated atherosclerotic potential (Alonso et al., Journal of American College of Cardiology Vol. 63, No. 19, 2014). In terms of diagnosis and preventive medicine, a patient's serum Lp(a) level is an independent and highly prevalent genetic risk factor for coronary heart disease and aortic stenosis (Saeedi and Frohlich Clinical Diabetes and Endocrinology (2016) 2:7). Currently, there is no approved specific Lp(a) reduction therapy beyond standard indirect LDL reduction measures. Consequently, methods are currently needed for the effective treatment, prevention, and risk reduction of suffering from disorders similar to and associated with stroke, atherosclerosis, thrombosis, and cardiovascular diseases such as coronary heart disease, aortic stenosis, and other as-yet-unidentified associated disorders, pathologies, or syndromes. The present invention addresses this unmet medical need. SUMMARY OF THE INVENTION [005] One aspect of the invention relates to a nucleic acid for inhibiting LPA expression in a cell, comprising at least one duplex region comprising at least one portion of a first strand and at least one portion of a second strand that is at least partially complementary to the first strand, wherein said first strand is at least partially complementary to at least one portion of RNA transcribed from the LPA gene, wherein said first strand comprises a nucleotide sequence selected from the following sequences: SEQ ID NOs: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, [006] wherein the nucleotides at positions 2 and 14 of the 5' end of the first strand are modified with a fluoro 2' modification and the nucleotides in the second strand that correspond to positions 11-13 of the first strand are modified with a fluoro 2' modification. [007] The invention also provides a composition comprising the nucleic acid or conjugated nucleic acid of any aspect of the invention, and optionally a physiologically acceptable excipient. [008] One aspect refers to a nucleic acid that is capable of inhibiting LPA expression for use as a drug. [009] A nucleic acid or conjugated nucleic acid is al