A siRNA targeting ANGPTL4 and its conjugate and use

Abstract

The present invention provides an siRNA for inhibiting the expression of angiopoietin-like protein 4 (ANGPTL4) gene, which contains a sense strand and an antisense strand, wherein the antisense strand contains at least 17 consecutive nucleotides that differ by no more than 4 nucleotides from the nucleotide sequence shown in any one of SEQ ID NO:2 to SEQ ID NO:265, and the antisense strand is 17 to 30 nucleotides in length; the sense strand is 17 to 30 nucleotides in length and is at least partially complementary to the antisense strand. The siRNA, siRNA conjugate and pharmaceutical composition provided by the present invention have good stability, excellent ANGPTL4 gene inhibition activity, satisfactory cytotoxicity and immunostimulation, and can significantly reduce the ANGPTL4 protein concentration at the animal level.

Description

Technical Field

[0001] The present invention relates to an siRNA for inhibiting the expression of angiopoietin-like protein 4 (ANGPTL4) gene, a conjugate thereof, a pharmaceutical composition and use thereof in preventing and / or treating diseases related to dyslipidemia. Background Art

[0002] Angiopoetin-like 4 (ANGPTL4) is a member of the angiopoietin-like protein family. It is a secretory protein that is mainly expressed in adipose and liver tissues and then secreted into the blood. It contains 406 amino acid sequences, mainly including two functional domains: the N-terminal coiled-coil domain and the C-terminal fibrinogen-like domain. It often exists in the form of oligomers, glycosylation, and various subtypes; and both domains are conserved domains of angiopoietin family members. In addition, ANGPTL4 also contains an asparagine glycosylation site, a cAMP, cGMP-dependent protein kinase phosphorylation site, 2 protein kinase C phosphorylation sites, 4 myristoylation sites, and 4 casein kinase II phosphorylation sites. Its expression is regulated by factors such as transforming growth factor-β (TGF-β), peroxisome proliferator-activated receptorδ (PPARδ), and hypoxia-inducible factor 1alpha (HIF1α).

[0003] At present, more and more evidence shows that the biological function of ANGPTL4 plays a key role in the pathological changes of metabolic diseases such as atherosclerosis, type 2 diabetes, fatty liver, and obesity. In particular, it can play a key role in lipid metabolism in the blood and liver by inhibiting the activity of LPL enzyme. Abnormal lipid metabolism in the liver can lead to the accumulation of TAG (Triacylglycerol) and DAG (1,2-diacylglycerol), thereby activating the PKCε signaling pathway, resulting in the inhibition of insulin receptor activation and reduced liver insulin sensitivity. Reduced liver insulin sensitivity inhibits the conversion of blood sugar into glycogen, promotes the dissimilation of glycogen into glucose, and leads to increased blood sugar. Several studies have shown that the expression of ANGPTL4 is related to lipid and carbohydrate metabolism in organisms.

[0004] Type 2 diabetes and its related complications have a high incidence rate worldwide. According to statistics from the International Diabetes Federation, there are currently about 537 million adults with diabetes (10%), and this number is expected to rise to 643 million by 2023 and 783 million by 2045, of which 90% are type 2 diabetes. In my country, according to the "Guidelines for the Prevention and Treatment of Type 2 Diabetes in China 2020 Edition", the incidence of type 2 diabetes in my country has risen to 11.2%. Although drugs for the treatment of type 2 diabetes have been launched in recent years, clinical blood sugar control and patient compliance are still not ideal. A survey published in the British Medical Journal in 2020 showed that the treatment rate of type 2 diabetes in China was 49%, and the proportion of patients with glycated hemoglobin (HbA1c) reaching the standard was only 49.4%. At the same time, the frequency of administration of existing drugs for the treatment of type 2 diabetes is high, and most of them need to be taken every day or even every meal, and patient compliance needs to be improved.

[0005] The present application provides a small interfering RNA (siRNA) preparation targeting ANGPTL4, which can destroy the normal translation template function of ANGPTL4 mRNA by specifically binding to ANGPTL4 mRNA, thereby preventing it from translating ANGPLT4 protein, thereby relieving the inhibition of LPL enzyme activity, reducing the content of TAG and DAG, reducing the PKCε signaling pathway, improving insulin sensitivity, and improving blood lipid and blood sugar metabolism.

[0006] Compared with traditional drugs, siRNA has poor stability and is easily degraded by nucleases when administered systemically. In addition, it is necessary to try to further improve the activity while avoiding side effects such as off-target effects, immune stimulation, and cytotoxicity. Therefore, it is urgent to develop more candidate siRNAs that are stable in the blood, have good biological activity, low cytotoxicity, and can inhibit ANGPTL4 gene expression for a long time. At the same time, there is a need for clinical research and commercialization of drugs that can effectively prevent and / or treat diseases related to abnormal blood lipid and blood sugar metabolism using the above candidate siRNAs that inhibit ANGPTL4 gene expression. Summary of the invention

[0007] The present invention provides an siRNA for inhibiting ANGPTL4 gene expression, characterized in that the siRNA comprises a sense strand and an antisense strand, wherein the antisense strand comprises at least 17 consecutive nucleotides that differ by no more than 4 nucleotides from the nucleotide sequence shown in any one of SEQ ID NO:2 to SEQ ID NO:265, and the antisense strand is 17 to 30 nucleotides in length; the sense strand is 17 to 30 nucleotides in length and is at least partially complementary to the antisense strand.

[0008] In some embodiments of the present invention, the antisense strand is 19 to 27 nucleotides in length; and the sense strand is 19 to 25 nucleotides in length.

[0009] In some embodiments of the present invention, the antisense strand is 19 to 23 nucleotides in length; and the sense strand is 19 to 21 nucleotides in length.

[0010] In some embodiments of the invention, the antisense strand is 23 nucleotides in length and the sense strand is 21 nucleotides in length. In some embodiments of the invention, the antisense strand is 21 nucleotides in length and the sense strand is 21 nucleotides in length. In some embodiments of the invention, the antisense strand is 21 nucleotides in length and the sense strand is 19 nucleotides in length. In some embodiments of the invention, the antisense strand is 19 nucleotides in length and the sense strand is 19 nucleotides in length.

[0011] In some embodiments of the invention, the antisense strand is 27 nucleotides in length and the sense strand is 25 nucleotides in length.

[0012] In some embodiments of the invention, the siRNA comprises one or more single-stranded nucleotide overhangs. For example, an overhang of 1, 2, 3 or 4 nucleotides. In some embodiments of the invention, the overhang can be on the sense strand, the antisense strand or any combination thereof. In some embodiments of the invention, the overhang is present at the 5' end, the 3' end or both ends of the siRNA antisense strand or sense strand.

[0013] In some embodiments of the present invention, the 3' end of the antisense strand of the siRNA has an overhang of 2 nucleotides.

[0014] In some embodiments of the present invention, the 3' end of the antisense strand of the siRNA has an overhang of 2 nucleotides, and the overhang is GG.

[0015] In some embodiments of the invention, the siRNA has blunt ends. In some embodiments of the invention, the siRNA has at least one blunt end located at the 5' end of the antisense strand (or the 3' end of the sense strand).

[0016] In some embodiments of the invention, the siRNA has two blunt ends.

[0017] In some embodiments of the present invention, the antisense strand differs from any nucleotide sequence shown in SEQ ID NO:2 to SEQ ID NO:265 by no more than 4 nucleotides; in some embodiments of the present invention, the antisense strand differs from any nucleotide sequence shown in SEQ ID NO:2 to SEQ ID NO:265 by no more than 3 nucleotides; in some embodiments of the present invention, the antisense strand differs from any nucleotide sequence shown in SEQ ID NO:2 to SEQ ID NO:265 by no more than 2 nucleotides; in some embodiments of the present invention, the antisense strand differs from any nucleotide sequence shown in SEQ ID NO:2 to SEQ ID NO:265 by no more than 1 nucleotide; in some embodiments of the present invention, the antisense strand is any nucleotide sequence shown in SEQ ID NO:2 to SEQ ID NO:265.

[0018] In some embodiments of the present invention, the sense strand and the antisense strand have no more than 3 nucleotide mismatches; in some embodiments of the present invention, the sense strand and the antisense strand have no more than 2 nucleotide mismatches; in some embodiments of the present invention, the sense strand and the antisense strand have no more than 1 nucleotide mismatch; in some embodiments of the present invention, the sense strand and the antisense strand are fully complementary.

[0019] In some embodiments of the present invention, the siRNA is selected from duplex 1 to duplex 264.

[0020] In some embodiments of the present invention, the siRNA contains at least one modified nucleotide.

[0021] In some embodiments of the present invention, all nucleotides in the sense strand and / or antisense strand of the siRNA are modified nucleotides or nucleotide analogs.

[0022] In some embodiments of the present invention, the siRNA is selected from modified duplex 1 to modified duplex 21.

[0023] The present invention also provides a siRNA conjugate obtained by conjugating any of the above siRNAs with a conjugation molecule.

[0024] In some embodiments of the present invention, the siRNA conjugate is selected from Conjugate 1 to Conjugate 72.

[0025] The present invention also provides a pharmaceutical composition, comprising any of the above siRNAs and / or any of the above siRNA conjugates and a pharmaceutically acceptable carrier.

[0026] The present invention also provides use of any of the above siRNAs and / or any of the above siRNA conjugates and / or the above pharmaceutical compositions in the preparation of a medicament for treating and / or preventing pathological conditions or diseases associated with overexpression of the angiopoietin-like protein 4 (ANGPTL4) gene.

[0027] Furthermore, the pathological condition or disease is a disease associated with dyslipidemia; further preferably, the disease associated with dyslipidemia is hyperlipidemia, hypertriglyceridemia, pancreatitis, familial chylomicronemia syndrome, diabetes, type 2 diabetes, heart disease, myocardial infarction, angina pectoris or atherosclerosis.

[0028] The siRNA, siRNA conjugates and pharmaceutical compositions provided by the present disclosure have good stability, excellent ANGPTL4 gene inhibition activity, satisfactory cytotoxicity and immunostimulation, and can significantly reduce blood lipid levels.

[0029] The sequence of the ANGPTL4 gene targeted by the siRNA in the present invention is shown in SEQ ID NO: 1:

[0030] SEQ ID NO: 1 (ANGPTL4 gene)

[0031] 1agaagccgag ctgagcggat cctcacacga ctgtgatccg attctttcca gcggcttctg

[0032] 61caaccaagcg ggtcttaccc ccggtcctcc gcgtctccag tcctcgcacc tggaacccca

[0033] 121acgtccccga gagtccccga atccccgctc ccaggctacc taagaggatg agcggtgctc

[0034] 181cgacggccgg ggcagccctg atgctctgcg ccgccaccgc cgtgctactg agcgctcagg

[0035] 241gcggacccgt gcagtccaag tcgccgcgct ttgcgtcctg ggacgagatg aatgtcctgg

[0036] 301cgcacggact cctgcagctc ggccaggggc tgcgcgaaca cgcggagcgc acccgcagtc

[0037] 361agctgagcgc gctggagcgg cgcctgagcg cgtgcgggtc cgcctgtcag ggaaccgagg

[0038] 421ggtccaccga cctcccgtta gcccctgaga gccgggtgga ccctgaggtc cttcacagcc

[0039] 481tgcagacaca actcaaggct cagaacagca ggatccagca actcttccac aaggtggccc

[0040] 541agcagcagcg gcacctggag aagcagcacc tgcgaattca gcatctgcaa agccagtttg

[0041] 601gcctcctgga ccacaagcac ctagaccatg aggtggccaa gcctgcccga agaaagaggc

[0042] 661tgcccgagat ggcccagcca gttgacccgg ctcacaatgt cagccgcctg caccggctgc

[0043] 721ccagggattg ccaggagctg ttccaggttg gggagaggca gagtggacta tttgaaatcc

[0044] 781agcctcaggg gtctccgcca tttttggtga actgcaagat gacctcagat ggaggctgga

[0045] 841cagtaattca gaggcgccac gatggctcag tggacttcaa ccggccctgg gaagcctaca

[0046] 901 aggcggggtt tggggatccc cacggcgagt tctggctggg tctggagaag gtgcatagca

[0047] 961tcacggggga ccgcaacagc cgcctggccg tgcagctgcg ggactgggat ggcaacgccg

[0048] 1021agttgctgca gttctccgtg cacctgggtg gcgaggacac ggcctatagc ctgcagctca

[0049] 1081ctgcacccgt ggccggccag ctgggcgcca ccaccgtccc acccagcggc ctctccgtac

[0050] 1141ccttctccac ttgggaccag gatcacgacc tccgcaggga caagaactgc gccaagagcc

[0051] 1201tctctggagg ctggtggttt ggcacctgca gccattccaa cctcaacggc footcttcc

[0052] 1261 gctccatccc acagcagcgg cagaagctta agaaggggaat cttctggaag acctggcggg

[0053] 1321gccgctacta cccgctgcag gccaccacca tgttgatcca gcccatggca gcagaggcag

[0054] 1381cctcctagcg tcctggctgg gcctggtccc aggcccacga aagacggtga ctcttggctc

[0055] 1441tgcccgagga tgtggccgtt ccctgcctgg gcaggggctc caaggagggg ccatctggaa

[0056] 1501acttgtggac agagaagaag accacgactg gagaagcccc ctttctgagt gcaggggggc

[0057] 1561tgcatgcgtt gcctcctgag atcgaggctg caggatatgc tcagactcta gaggcgtgga

[0058] 1621ccaaggggca tggagcttca ctccttgctg gccagggagt tggggactca gagggaccac

[0059] 1681ttggggccag ccagactggc ctcaatggcg gactcagtca cattgactga cggggaccag

[0060] 1741ggcttgtgtg ggtcgagagc gccctcatgg tgctggtgct gttgtgtgta ggtcccctgg

[0061] 1801ggacacaagc aggcgccaat ggtatctggg cggagctcac agagttcttg gaataaaagc

[0062] 1861aacctcagaa ca

[0063] In the present invention, unless otherwise specified, capital letters C, G, U, and A represent base compositions of modified or unmodified nucleotides; lowercase letter d represents that a nucleotide adjacent to the right side of the letter d is a deoxyribonucleotide; lowercase letter m represents that a nucleotide adjacent to the right side of the letter m is a 2'-methoxy-modified nucleotide; mark f represents that a nucleotide adjacent to the right side of the mark f is a 2'-fluorine-modified nucleotide; mark * represents that two nucleotides adjacent to the left and right of the mark * are connected by thiophosphate groups; and the letters eVP represent that a nucleotide adjacent to the right side of the mark eVP is a vinyl phosphate-modified nucleotide.

[0064] In the present invention, unless otherwise specified, the term "complementary" refers to the ability of an oligonucleotide of a first sequence to hybridize with an oligonucleotide of a second sequence under certain conditions and form a double-stranded structure. "At least partially complementary" means that the two sequences may be completely complementary, or generally no more than 5, 4, 3 or 2 mismatched base pairs, while retaining the ability to hybridize under relevant conditions. In addition, in the case where two oligonucleotides are designed to form one or more single-stranded overhangs when hybridized, such overhangs should not be regarded as mismatches in terms of determining complementarity. In the present invention, when the above hybridization ability requirements are met, the "complementary" sequence may also include or be completely formed from non-Watson-Crick base pairs and / or base pairs formed from non-natural and modified nucleotides. Such non-Watson-Crick base pairs include, but are not limited to, G:U wobble base pairing or Hoogstein base pairing. Correspondingly, in the present invention, unless otherwise specified, "mismatch" means that in the siRNA duplex molecule, the bases at corresponding positions are not paired in a complementary form.

[0065] In the present invention, unless otherwise specified, "difference in nucleotide sequence" means that the base type of the nucleotide at the same or corresponding position has changed compared to the original nucleotide sequence. For example, when one nucleotide base in the original nucleotide sequence is A, and the nucleotide base at the same or corresponding position is changed to U, C, G or dT, dC, dG, etc., it is considered that there is a difference in nucleotide sequence at this position. It should be noted here that, when the nucleotide at the same or corresponding position differs only in the presence or absence of modification or the type of modification compared to the original nucleotide sequence, it is not considered that there is a difference in nucleotide sequence at this position.

[0066] In the present invention, unless otherwise specified, the term "pharmaceutically acceptable" means that the carrier, vehicle, diluent, excipient and / or the salt / ester / hydrate formed therefrom are usually chemically or physically compatible with other ingredients constituting a drug dosage form and physiologically compatible with the receptor.

[0067] In the present invention, unless otherwise specified, the term "inhibition" refers to a situation where the expression of a target gene is down-regulated due to siRNA-mediated degradation of the mRNA of the target gene. The "down-regulation" refers to a situation where the expression level of the target gene decreases by 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 99% or more or even 100% relative to the situation without siRNA treatment. Among them, a 100% decrease in the expression level of the target gene means that there is no detectable level of target gene expression.

[0068] In some embodiments of the present invention, the siRNA may further contain modified nucleotides as needed, and the modified nucleotides will not significantly weaken or lose the function of the siRNA in inhibiting the expression of the ANGPTL4 gene. Currently, there are many ways to modify siRNA in the art, including, for example, backbone modification (such as phosphate group modification), ribose group modification, and base modification (Watts, JK, GF Deleavey, and MJ Damha, Chemically modified siRNA: tools and applications. Drug Discov Today, 2008. 13 (19-20): p. 842-55).

[0069] In some embodiments of the present invention, at least one nucleotide in the sense strand or antisense strand of the siRNA is a modified nucleotide, for example, the modified nucleotide is a nucleotide group in which the ribose group and optionally the phosphate group are modified, but is not limited thereto.

[0070] In some embodiments of the present invention, all nucleotides in the sense strand and / or the antisense strand are modified nucleotides or nucleotide analogs.

[0071] In some embodiments of the present invention, the modified nucleotide is selected from: 2'-methoxy nucleotides, 2'-fluoro nucleotides, 2'-deoxy nucleotides, 2',3'-split ring nucleotide analogs, 2'-fluoroarabino nucleotides, 2'-methoxyethyl nucleotides, 2'-amino modified nucleotides, 2'-alkyl modified nucleotides, 3'-methoxy nucleotides, 2'-allyl modified nucleotides, nucleotides containing thiophosphate groups, nucleotides containing methylphosphonate groups, nucleotides containing 5'-phosphate esters, nucleotides containing 5'-phosphate mimetics, diol-modified nucleotides, abasic nucleotides, morpholino nucleotides, locked nucleotides, unlocked nucleotides or glycerol nucleotides, but the present invention is not limited thereto.

[0072] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 6th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0073] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 6th, 8th, 9th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0074] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 6th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0075] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 6th, 8th, 9th, 10th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0076] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 6th, 8th, 9th, 10th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, 11th, and 15th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0077] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 6th, 10th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 3rd, 7th, 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0078] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 6th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0079] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 3rd, 4th, 6th, 8th, 10th, 14th, 16th, 18th, 20th, and 22nd positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 2nd position at the 5' end of the sense strand is a 2'-fluoro nucleotide, and the remaining positions are 2'-methoxy nucleotides.

[0080] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 4th, 5th, 6th, 8th, 10th, 14th, 16th, 18th, 20th, and 22nd positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 2nd position at the 5' end of the sense strand is a 2'-fluoro nucleotide, and the remaining positions are 2'-methoxy nucleotides.

[0081] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 4th, 6th, 7th, 8th, 10th, 14th, 16th, 18th, 20th, and 22nd positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 2nd position at the 5' end of the sense strand is a 2'-fluoro nucleotide, and the remaining positions are 2'-methoxy nucleotides.

[0082] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 4th, 6th, 8th, 10th, 14th, 16th, 18th, and 20th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 1st, 3rd, 5th, 7th, 9th, 10th, 11th, 13th, 15th, 17th, 19th, and 21st positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0083] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 4th, 6th, 8th, 10th, 14th, 16th, 18th, 20th, and 22nd positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 1st, 3rd, 5th, 7th, 9th, 10th, 11th, 13th, 15th, 17th, 19th, and 21st positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0084] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 6th, 8th, 9th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, the 7th position is a glycerol nucleotide (GNA), and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0085] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 8th, 9th, 10th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, the 6th position is a glycerol nucleotide (GNA), and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, 11th, and 15th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0086] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 8th, 9th, 10th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, the 4th position is a glycerol nucleotide (GNA), and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, 11th, and 15th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0087] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 8th, 9th, 10th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, the 5th position is a glycerol nucleotide (GNA), and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, 11th, and 15th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0088] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 8th, 9th, 10th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, the 7th position is a glycerol nucleotide (GNA), and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, 11th, and 15th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0089] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 8th, 9th, 10th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, the 6th position is a glycerol nucleotide (GNA), and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0090] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 5th, 7th, and 12th positions of the 5' end of the antisense strand are 2'-deoxynucleotides, the 14th position is a 2'-fluoro nucleotide, and the remaining positions are 2'-methoxy nucleotides. In some embodiments of the present invention, the sense strand of the siRNA is 21 nucleotides in length, wherein the 9th, 10th, and 11th positions of the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0091] In some embodiments of the present invention, the antisense strand of the siRNA is 23 nucleotides in length, wherein the 2nd, 5th, 7th, and 12th positions of the 5' end of the antisense strand are 2'-deoxynucleotides, the 6th, 8th, 9th, 10th, 14th, and 16th positions are 2'-fluoronucleotides, and the remaining positions are 2'-methoxynucleotides. In some embodiments of the present invention, the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, and 11th positions of the 5' end of the sense strand are 2'-fluoronucleotides, and the remaining positions are 2'-methoxynucleotides.

[0092] In some embodiments of the present invention, the antisense strand of the siRNA is 21 nucleotides in length, wherein the 2nd, 6th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 5th, 7th, and 9th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0093] In some embodiments of the present invention, the antisense strand of the siRNA is 21 nucleotides in length, wherein the 2nd, 6th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 7th, 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0094] In some embodiments of the present invention, the antisense strand of the siRNA is 21 nucleotides in length, wherein the 2nd, 4th, 6th, 8th, 10th, 12th, 14th, 16th, 18th, and 20th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 9th, 10th, and 11th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0095] In some embodiments of the present invention, the antisense strand of the siRNA is 21 nucleotides in length, wherein the 2nd, 4th, 6th, 8th, 10th, 12th, 14th, 16th, 18th, and 20th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 21 nucleotides in length, wherein the 1st, 3rd, 5th, 7th, 9th, 11th, 13th, 15th, 17th, 19th, and 21st positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0096] In some embodiments of the present invention, the antisense strand of the siRNA is 21 nucleotides in length, wherein the 2nd, 6th, 14th, and 16th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 19 nucleotides in length, wherein the 7th, 8th, and 9th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0097] In some embodiments of the present invention, the antisense strand of the siRNA is 19 nucleotides in length, wherein the 2nd, 4th, 6th, 8th, 10th, 12th, 14th, 16th, and 18th positions at the 5' end of the antisense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides; the sense strand of the siRNA is 19 nucleotides in length, wherein the 7th, 8th, and 9th positions at the 5' end of the sense strand are 2'-fluoro nucleotides, and the remaining positions are 2'-methoxy nucleotides.

[0098] In some embodiments of the present invention, the modified nucleotide is a nucleotide in which the phosphate group is modified by a phosphorothioate group, that is, a non-bridging oxygen atom in a phosphodiester bond is replaced by a sulfur atom, thereby replacing the phosphodiester bond with a phosphorothioate diester bond.

[0099] In some embodiments of the present invention, at least one of the nucleotides between the 1st and 2nd positions at the 5' end of the sense chain, between the 2nd and 3rd positions at the 5' end of the sense chain, between the 1st and 2nd positions at the 3' end of the sense chain, between the 2nd and 3rd positions at the 3' end of the sense chain, between the 1st and 2nd positions at the 3' end of the antisense chain, between the 2nd and 3rd positions at the 3' end of the antisense chain, between the 1st and 2nd positions at the 5' end of the antisense chain, and between the 2nd and 3rd positions at the 5' end of the antisense chain is connected by thiophosphate groups; preferably, at least four of them are connected by thiophosphate groups; in some embodiments of the present invention, at least six of them are connected by thiophosphate groups; in some embodiments of the present invention, all eight of them are connected by thiophosphate groups.

[0100] In some embodiments of the present invention, the nucleotides at the first and second positions and the nucleotides at the second and third positions at the 5' end of the sense strand are linked by phosphorothioate groups.

[0101] In some embodiments of the present invention, the 1st and 2nd nucleotides and the 2nd and 3rd nucleotides at the 5' end of the sense strand are linked by phosphorothioate groups, and the 1st and 2nd nucleotides and the 2nd and 3rd nucleotides at the 3' end are linked by phosphorothioate groups.

[0102] In some embodiments of the present invention, the nucleotides at positions 1 and 2 and 2 and 3 at the 3' end of the antisense strand are linked by phosphorothioate groups, and the nucleotides at positions 1 and 2 and 2 and 3 at the 5' end are linked by phosphorothioate groups.

[0103] In some embodiments of the invention, the sense strand may include one or more blocking residues or moieties, referred to as "blocking residues". A "blocking residue" is a non-nucleotide compound or other moiety that can be incorporated into one or more ends of the nucleotide sequence of the siRNA. In some embodiments of the invention, the blocking residue is present at the 5' end, the 3' end, or both the 5' end and the 3' end of the sense strand.

[0104] In some embodiments of the present invention, an inverted abasic residue (iab) is added as a capping residue. See F. Czauderna, Nucleic Acids Res., 2003, 31(11), 2705-16. In some embodiments of the present invention, the 5' end and / or 3' end of the sense strand may contain more than one inverted abasic deoxyribose moiety as a capping residue.

[0105] In some embodiments of the invention, one or more reverse abasic residues (iab) are added to the 3' end of the sense strand. In some embodiments of the invention, one or more reverse abasic residues (iab) are added to the 5' end of the sense strand. In some embodiments of the invention, one or more reverse abasic residues may be inserted between the linker-targeting ligand portion and the nucleotide sequence of the siRNA sense strand. In some embodiments of the invention, one or more reverse abasic residues are included at or near one or more ends of the siRNA sense strand.

[0106] In some embodiments of the invention, one or more inverted abasic residues (iab) are added to the 5' end of the sense strand. In some embodiments of the invention, one or more inverted abasic residues may be inserted between the linker-targeting ligand portion and the nucleotide sequence of the siRNA sense strand.

[0107] Inverted abasic residues may be linked via phosphate, phosphorothioate or other internucleoside linkages.

[0108] In some embodiments of the present invention, the first nucleotide at the 5' end of the antisense strand is selected from the following structures:

[0109]

[0110] In some embodiments of the present invention, the first nucleotide at the 5' end of the antisense strand is selected from the following structures:

[0111]

[0112] In some embodiments of the present invention, the first nucleotide at the 5' end of the antisense strand is a nucleotide modified with (E)-vinyl phosphate.

[0113] In the present invention, unless otherwise specified, "conjugation" refers to the connection between two or more chemical moieties by covalent bonding; "conjugate" refers to a compound formed by covalent bonding between chemical moieties; "siRNA conjugate" refers to a compound formed by covalent bonding of one or more chemical moieties to siRNA. It should be noted that each chemical moiety can be directly bonded to siRNA or can be bonded to siRNA through a linker.

[0114] In the present invention, unless otherwise specified, the “-” in “linker-targeting ligand” means that the linker and the targeting ligand are covalently linked.

[0115] In some embodiments of the present invention, the siRNA of the present invention can be conjugated with a pharmaceutically acceptable conjugated molecule to obtain a siRNA conjugate. In some embodiments of the present invention, the siRNA is covalently conjugated to the conjugated molecule. In order to reduce the effect that the conjugation may have on the activity of the siRNA, the conjugation site of the siRNA and the conjugated molecule can be at the 3' end or 5' end of the siRNA sense strand, or at the 5' end of the antisense strand. In some embodiments, the conjugation site of the siRNA and the conjugated molecule can also be in the internal sequence of the siRNA.

[0116] The pharmaceutically acceptable targeting ligand can be a targeting ligand conventionally used in the field of siRNA administration, such as, but not limited to, one or more of the following targeting ligands or their derivatives: lipophilic molecules, such as cholesterol, bile acid, vitamins (such as vitamin E), lipid molecules of different chain lengths; polymers, such as polyethylene glycol; polypeptides, such as membrane-permeable peptides; aptamers; antibodies; quantum dots; carbohydrates, such as lactose, polylactose, mannose, galactose, N-acetylgalactosamine (GalNAc); folic acid (folate); or receptor ligands expressed by hepatocytes, such as asialoglycoproteins, asialosugar residues, lipoproteins (such as high-density lipoproteins, low-density lipoproteins, etc.), glucagon, neurotransmitters (such as adrenaline), growth factors, transferrin, etc.

[0117] In some embodiments of the invention, the targeting ligand is N-acetylgalactosamine.

[0118] In some embodiments of the invention, the targeting ligand is directly linked to the 3' end of the sense strand of the siRNA.

[0119] In some embodiments of the invention, the targeting ligand is directly linked to the 5' end of the sense strand of the siRNA.

[0120] In some embodiments of the invention, the targeting ligand is linked to the 3' end of the sense strand of the siRNA via a linker. In some embodiments of the invention, the targeting ligand is linked to the 5' end of the sense strand of the siRNA via a linker.

[0121] In some embodiments of the invention, the Linker-Targeting Ligand portion has the following structure:

[0122]

[0123] Wherein, R is selected from hydrogen and auxiliary groups. In some embodiments of the present invention, the auxiliary groups include but are not limited to long-chain alkyl groups, long-chain alkenyl groups, long-chain alkynyl groups, cholesterol groups, cholesterol-like groups, and polyethylene glycol groups; in some embodiments of the present invention, the auxiliary groups include but are not limited to polypeptides.

[0124] In some embodiments of the invention, the linker-targeting ligand portion is GalNAc(L96) having the following structure:

[0125]

[0126] In some embodiments of the invention, GalNAc(L96) is linked to the 3' end of the sense strand of the siRNA.

[0127] In some embodiments of the invention, GalNAc(L96) is attached to the inverted abasic residue (iab) at the 3' end of the sense strand of the siRNA.

[0128] In some embodiments of the invention, the linker-targeting ligand portion is Ser(GN) having the following structure:

[0129] In some embodiments of the invention, Ser(GN) is linked to the 3' end of the sense strand of the siRNA. In some embodiments of the invention, Ser(GN) is linked to the 5' end of the sense strand of the siRNA.

[0130] In some embodiments of the invention, Ser(GN) is attached to both the 3' end and the 5' end of the sense strand of the siRNA.

[0131] The present invention also provides a pharmaceutical composition, which contains the siRNA disclosed in the present invention as an active ingredient and a pharmaceutically acceptable carrier.

[0132] In some embodiments of the present disclosure, the pharmaceutical composition contains one siRNA as described in the first aspect. In other embodiments of the present disclosure, the pharmaceutical composition contains at least two siRNAs as described in the first aspect (for example, but not limited to 2, 3, 4, 5, 6, 7, 8, 9, 10 or more) as active ingredients. Preferably, the at least two siRNAs as described in the first aspect each target a different target sequence in the ANGPTL4 gene, thereby being expected to act simultaneously against different target sequences to bring about a synergistic effect. Here, the so-called "different target sequences" means that there is no overlap between the target sequences, or the number of overlapping continuous nucleotides between the target sequences is less than 5 (for example, the number of overlapping continuous nucleotides is 4, 3, 2, 1, 0). In this case, the at least two siRNAs as described in the first aspect may be present in any different ratios. Preferably, the at least two siRNAs as described in the first aspect may be present in a molar ratio of 1:100 to 100:1; more preferably, the at least two siRNAs as described in the first aspect may be present in a molar ratio of 1:10 to 10:1, 1:5 to 5:1 or 1:2 to 2:1. In some embodiments of the present disclosure, the at least two siRNAs as described in the first aspect are present in the same molar ratio. DETAILED DESCRIPTION

[0133] It is known to those skilled in the art that the siRNA of the present invention can be obtained by conventional siRNA preparation methods in the art (e.g., solid phase synthesis and liquid phase synthesis), wherein both solid phase synthesis and liquid phase synthesis have commercial customization services. It is also clear to those skilled in the art that modified nucleotide groups can be introduced into the siRNA of the present invention by using nucleotide monomers with corresponding modifications. Methods for preparing nucleotide monomers with corresponding modifications are well known to those skilled in the art, and commercial monomers are also available on the market.

[0134] Example 1: siRNA Synthesis

[0135] For the sense and antisense strands of the siRNA sequence of the present invention and the sense and antisense strands of the modified duplex, deoxynucleoside CPG is used as a solid support; the sense strand is synthesized using the solid support, and the antisense strand is synthesized using universal CPG.

[0136] The sequence was synthesized on a 48-channel synthesizer at a scale of 0.2 μmol. The phosphoramidite monomer was used at a concentration of 0.05 M and the activator was 0.3 M BTT.

[0137] The cutting and deprotection of the sequence were carried out in a 1.5 ml tube, with AMA used in the first step and triethylamine trihydrofluoride used in the second step to remove the two protecting groups. For sequences containing two fully modified sequences, ammonia solution was required. Acetone: ethanol (80:20) mixture was used to precipitate the sequence after cutting and deprotection and to dissolve it in RNase-free water. Each sequence was analyzed by LC-MS to determine sequence accuracy, quantitatively by spectrophotometer and purity was determined by HPLC.

[0138] After HPLC purification, freeze-drying and quality inspection, the salt was replaced by sodium acetate alcohol precipitation and desalted using a 3KD ultrafiltration tube. After desalting, the sense chain and antisense chain were quantitatively determined by a spectrophotometer and mixed in a 1:1 ratio and annealed to form a siRNA duplex.

[0139] Table 1. Sense and antisense strand sequences of unmodified siRNA duplexes

[0140]

[0141]

[0142]

[0143]

[0144]

[0145]

[0146] Table 2. Sense and antisense strand sequences of modified siRNA duplexes

[0147]

[0148]

[0149] Part of the siRNA sequence was modified and conjugated to the linker-targeting ligand portion, and the siRNA conjugate was as follows:

[0150] Table 3. Sequences of modified siRNA conjugates

[0151]

[0152]

[0153]

[0154]

[0155]

[0156]

[0157] The effects of the present invention are described below using test examples.

[0158] Experimental Example 1: ANGPTL4 siRNA in vitro activity detection - psiCHECK

[0159] Human and nonhuman primate cross-reactive ANGPTL4 siRNA candidate sequences were screened by bioinformatics analysis, and 223 pairs of potential ANGPTL4 agents were synthesized. To screen the target siRNA, the human ANGPTL4 cDNA sequence (accession number NMs139314.3) was cloned from a commercially available mammalian expression vector (Oreene, Rockville, MD) into a commercially available reporter-based screening plasmid (PsiCheck 2 (Promega, Madison, WI)), which produces Renilla luciferase / ANGPTL4 fusion mRNA. PsiCHECK screening was performed in 293T cells (Nanjing Kebai). 293T cells were seeded in 96-well plates at 20,000 cells / well; 223 pairs of ANGPTL4 siRNA were transfected at two concentrations, 50 ng of ANGPTL4-psiCHECK 2 plasmid per well and 0.3 μL of lipofectamine 2000 per well. After 24 h of incubation at 37°C in a 5% CO2 environment, the ANGPTL4 gene knockout activity was determined using a dual-luciferase reporter assay system (Promega, E2920). Each duplex was tested for 3-4 independent transfections. Gene knockout was determined by measuring Renilla luciferase normalized to the level of constitutively expressed firefly luciferase (Tables 3A and 3B)

[0160] The results of the in vitro activity test of siRNA duplexes are shown below.

[0161] Table 3A, in vitro activity test results of duplexes (Batch 1)

[0162]

[0163] Table 3B, in vitro activity test results of duplexes (Batch 2)

[0164]

[0165]

[0166]

[0167] The above experimental data show that the siRNA of the present invention has good ANGPTL4 gene knockout activity.

[0168] Experimental Example 2: ANGPTL4 RNAi in vitro activity detection-qPCR

[0169] U138-MG cells (ATCC) were seeded into 24-well plates at 40,000 cells / well; transfection was performed after incubation at 37°C, 5% CO2 for 16 h. siRNA and lipofectamine RNAiMax (Invitrogen) were co-transfected; after incubation at 37°C, 5% CO2 for 24 h, RNA was extracted using MolPure Magnttic Tissue / Cell Total RNA Kit (Yisheng; cat: 18600ES60). cDNA synthesis was performed using the gDNA removal cDNA synthesis kit from Quanshijin (Beijing Quanshijin Biotechnology Co., Ltd.; Beijing; China Cat#AE311-03). The ΔΔCt assay was used in ABIQuantStudio TM 6 Real-time fluorescence PCR system was used for real-time fluorescence PCR. Each duplex was tested for 3-4 independent transfections; each transfection was tested in triplicate.

[0170] The in vitro activity test results of ANGPTL4 siRNA duplexes and conjugates are shown below. The experimental results are shown in the following table:

[0171] Table 4. In vitro activity test results of ANGPTL4 siRNA

[0172]

[0173]

[0174] Table 5. In vitro IC of ANGPTL4 siRNA 50 Test Results

[0175]

[0176] Table 6. In vitro activity test results of ANGPTL4 modified siRNA

[0177]

[0178] Table 7. In vitro activity test results of ANGPTL4 modified siRNA conjugates

[0179]

[0180]

[0181] Table 8. In vitro IC of ANGPTL4 siRNA 50 Test Results

[0182] Conjugate No. IC50(pM) 16 88.56 15 162.80 62 154.70 63 167.90 24 193.50 65 151.40 36 74.16 37 88.56

[0183] The above test data indicate that the siRNA duplex and conjugate of the present invention have excellent cell ANGPTL4 gene knockout activity and may become a new drug for targeted inhibition of ANGPTL4 gene expression.

Claims

1. A siRNA for inhibiting ANGPTL4 gene expression, characterized in that: The siRNA comprises a sense strand and an antisense strand; wherein the antisense strand comprises at least 17 consecutive nucleotides that differ by no more than 4 nucleotides from the nucleotide sequence shown in any one of SEQ ID NO:2 to SEQ ID NO:265, and the antisense strand is 17 to 30 nucleotides in length; the sense strand is 17 to 30 nucleotides in length and is at least partially complementary to the antisense strand.

2. The siRNA according to claim 1, characterized in that: The antisense strand is 19 to 27 nucleotides long; the sense strand is 19 to 25 nucleotides long.

3. The siRNA according to claim 1, characterized in that: The antisense strand differs from any nucleotide sequence shown in SEQ ID NO:2 to SEQ ID NO:265 by no more than 4 nucleotides.

4. The siRNA according to claim 1, characterized in that: The sense strand has no more than 3 nucleotide mismatches with the antisense strand.

5. The siRNA according to any one of claims 1 to 4, characterized in that: The siRNA is selected from duplex 1 to duplex 264.

6. The siRNA according to any one of claims 1 to 5, characterized in that: The siRNA contains at least one modified nucleotide.

7. The siRNA according to any one of claims 1 to 6, characterized in that: All nucleotides in the sense strand and / or antisense strand of the siRNA are modified nucleotides or nucleotide analogs.

8. An siRNA conjugate obtained by conjugating the siRNA according to any one of claims 1 to 7 with a conjugating molecule.

9. The siRNA conjugate according to claim 8, characterized in that: The siRNA conjugate is selected from conjugate 1 to conjugate 72.

10. A pharmaceutical composition, characterized in that: The pharmaceutical composition comprises the siRNA according to any one of claims 1 to 7 and / or the siRNA conjugate according to any one of claims 8 to 9 and a pharmaceutically acceptable carrier.

11. Use of the siRNA according to any one of claims 1 to 7 and / or the siRNA conjugate according to any one of claims 8 to 9 and / or the pharmaceutical composition according to claim 10 in the preparation of a medicament for treating and / or preventing a pathological condition or disease associated with overexpression of the angiopoietin-like protein 4 (ANGPTL4) gene.

12. The use according to claim 11, characterized in that: The pathological condition or disease is a disease associated with dyslipidemia; further preferably, the disease associated with dyslipidemia is hyperlipidemia, hypertriglyceridemia, pancreatitis, familial chylomicronemia syndrome, diabetes, type 2 diabetes, heart disease, myocardial infarction, angina pectoris or atherosclerosis.

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