EP2285962A2 - Nouveaux agents thérapeutiques pour le traitement de l'hépatite - Google Patents

Nouveaux agents thérapeutiques pour le traitement de l'hépatite

Info

Publication number
EP2285962A2
EP2285962A2 EP09745480A EP09745480A EP2285962A2 EP 2285962 A2 EP2285962 A2 EP 2285962A2 EP 09745480 A EP09745480 A EP 09745480A EP 09745480 A EP09745480 A EP 09745480A EP 2285962 A2 EP2285962 A2 EP 2285962A2
Authority
EP
European Patent Office
Prior art keywords
gene
hepatitis
nucleic acid
acid molecule
repressor
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP09745480A
Other languages
German (de)
English (en)
Inventor
Jörg Friedrich SCHLAAK
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Individual
Original Assignee
Individual
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Individual filed Critical Individual
Publication of EP2285962A2 publication Critical patent/EP2285962A2/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/70—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
    • C12Q1/701—Specific hybridization probes
    • C12Q1/706—Specific hybridization probes for hepatitis
    • C12Q1/707—Specific hybridization probes for hepatitis non-A, non-B Hepatitis, excluding hepatitis D
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A61P31/14—Antivirals for RNA viruses
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/14—Type of nucleic acid interfering N.A.

Definitions

  • the present invention relates to the use of substances, in particular in the form of inhibitors or repressors, which are capable of regulating the gene activity of a gene associated with the multiplication or replication of hepatitis viruses, in particular hepatitis C viruses, in the context of The field of diagnosis and therapy of hepatitis, in particular hepatitis type C.
  • the gene associated with the multiplication or replication of hepatitis C viruses is preferably the human interferon-stimulated gene ISG1.
  • the present invention relates to the use of a substance which inhibits or at least degrades the gene activity of a substance associated with the multiplication or replication of hepatitis viruses, in particular hepatitis C viruses, in particular hepatitis C viruses, in particular inhibitors or repressors, for the manufacture of a medicament or medicament for the prophylactic or curative treatment of hepatitis, in particular hepatitis type C.
  • the present invention relates to the use of an interferon-stimulated gene, in particular ISGl 5, for the discovery or provision of a medicament for the prophylactic or curative treatment of hepatitis, in particular hepatitis type C, and / or for the prediction of individual drug effects and / or for the prediction of side effects or the response of medicinal products.
  • an interferon-stimulated gene in particular ISGl 5
  • ISGl 5 for the discovery or provision of a medicament for the prophylactic or curative treatment of hepatitis, in particular hepatitis type C, and / or for the prediction of individual drug effects and / or for the prediction of side effects or the response of medicinal products.
  • the present invention relates to a method for the identification of substances, in particular inhibitors or repressors, which are the gene activity of an interferon-stimulated gene, in particular in connection with the multiplication or replication of hepatitis viruses, in particular hepatitis C viruses ISG 15, or a method for the identification of substances that regulate the activity of the corresponding gene products or gene-related products. Furthermore, the present invention relates to a Process for improving the pharmacological properties of these substances.
  • the present invention relates to a pharmaceutical composition which contains at least one pharmacologically active substance, in particular inhibitor or repressor, which is found on the basis of the inventive methods, the substance having the gene activity of one with the replication or replication of hepatitis C viruses influenced gene, in particular inhibits or inhibits;
  • the present invention relates to a pharmaceutical composition, preferably for the prophylactic or therapeutic treatment of hepatitis C diseases, which contains at least one siRNA in effective, especially pharmaceutically effective amounts.
  • HCV hepatitis C virus
  • the hepatitis C virus belongs to the family of Flaviviridae and is the only representative of the genus Hepaciviren.
  • the hepatitis C virus was originally assigned to the so-called NonA-NonB hepatitis viruses, until in 1989 succeeded in the sequencing of the viral genome.
  • Today, six genotypes are characterized, which in turn subdivide into subtypes. Infection with the virus takes place in most cases via transfusion of infected blood reserves, especially in the seventies, or via cannula puncture wounds, for example, to hospital staff. Other transmission options include unprotected intercourse and the use of common injections among drug addicts.
  • the acute infection is usually asymptomatic. In about 70 to 80% of cases, chronic liver infection then occurs which can lead to liver cirrhosis and hepatocellular carcinoma.
  • Interferon in particular ⁇ -interferon (synonymously also referred to as interferon-alpha or IFN- ⁇ , preferably pegylated IFN- ⁇ , as the hitherto most efficient therapy of chronic hepatitis type C), is used in the prior art. optionally in combination with the antiviral ribavirin. Depending on the genotype and other factors, this therapy will cure only in 50% to 90% of patients. One of the most common side effects of this therapy is IFN-induced major depression, which, in addition to deteriorating quality of life, may lead to discontinuation or even suicide. A vaccine against the hepatitis C virus has not yet been developed.
  • Interferons are low-molecular proteins and are counted among the cytokines.
  • Type I interferons occur in monomeric form and can be subdivided into superfamilies whose main difference is their origin.
  • IFN- ⁇ and IFN-ß account for most of type I interferons.
  • the group of ⁇ -interferons in turn, can be subdivided into a number of subtypes that are expressed by different genes. They are produced largely in leukocytes and fibroblasts, while IFN-ß is mainly produced by endothelial cells and fibroblasts.
  • IFN- ⁇ and IFN- ⁇ belong to type I interferons.
  • Type I interferons is induced by the recognition of viral as well as bacterial pathogenic patterns. They are secreted by infected cells and act both paracrine on neighboring cells and autocrine on the IFN-producing cell itself. Binding to the receptor triggers a signal cascade that ends in the induction of interferon-stimulated genes (ISGs).
  • ISGs interferon-stimulated genes
  • the enzymes E1, E2 and E3 are also up-regulated by type I interferons, leading to increased ISG-ylation and enhancing the interferon response.
  • Uspl ⁇ the protease that reverses ISGylation, - A -
  • interferons are used for the therapy of tumors, autoimmune diseases and viral infections (e.g., HBV and HCV) because of their properties and the ability to induce a variety of cellular defense mechanisms and to support the body's immune response.
  • autoimmune diseases and viral infections e.g., HBV and HCV
  • the use of pegylated IFN- ⁇ 2a or IFN- ⁇ 2b in combination with the nucleoside analog ribavirin (RBV) has in particular established itself in the therapy of hepatitis type C diseases or infections.
  • the response to therapy depends on the HCV genotype, viral load, age of the patient, already incurred liver damage, and co-infections.
  • genotype 2 and 3 genomes up to 80% of patients have a sustained virus decline after 24 weeks, whereas only genotypes of genotype 1 infections respond to treatment, with a continuing decline in the number of genotypes Viral load after 48 weeks at a body weight-dependent RBV dosage have.
  • the response to therapy can be divided into two phases. Early viral response (EVR), depending on the genotype and the dose of drugs used in the first phase, is followed by a slower, but sustained, decreased viral response (SVR). in the second phase. Investigations have shown that, if within the first 12 weeks after initiation of therapy no EVR with a decrease in viral load by at least 21og 1 o-phases, no sustained response is expected and the therapy can be stopped. Different expression patterns of ISGs in both peripheral blood mononuclear cells (PBMCs) and liver biopsies from patients also allow predictions of response to therapy to be made. A disadvantage of the aforementioned therapy of the prior art are thus sometimes strong side effects, especially as regards the formation of depression, and the fact that an effective therapeutic success - as described above - is not always guaranteed.
  • PBMCs peripheral blood mononuclear cells
  • an object of the present invention is to provide new and efficient therapy or treatment options for hepatitis diseases, in particular hepatitis C diseases, which have over the approaches known in the art, a higher efficacy with reduced side effect should.
  • a further object of the present invention is to provide a method for the determination of substances or substances as such, with which the gene expression or gene activity related in connection with the multiplication or replication of hepatitis C viruses Genes, in particular ISGl 5, can be reduced or prevented in order to at least reduce virus replication or replication.
  • a further object of the present invention is to provide a method by which specific genes can be identified, which are significantly involved in the multiplication or replication of hepatitis C virus, which are preferably human or interferon-stimulated genes.
  • the Applicant has succeeded, in a completely surprising manner, for the first time in identifying a specific gene which is significantly associated with the multiplication or replication of hepatitis C viruses in a host system, in particular in humans .
  • the Applicant has surprisingly found that the interferon-stimulated gene ISG 15 is significantly involved in the aforementioned proliferation or replication of hepatitis C virus and induction or activation of this gene can lead to an undesirable increase in virus replication.
  • the gene ISG1 5 identified with regard to its relevance with regard to the multiplication or replication of hepatitis C viruses can thus be used as a basis for methods or medicaments for the therapy of a hepatitis C disease or infection, according to the invention is turned off on an inactivation of the corresponding gene.
  • the Applicant has found in completely surprising manner that the specific use of an inhibitor or repressor, with which the (gene) activity of the gene in connection with the multiplication or replication of hepatitis C virus, in particular ISG 15, and thus influencing its gene activity leads to a significant reduction of hepatitis C viral load, which is largely due to a significantly reduced proliferation or replication of the hepatitis C virus.
  • the Applicant was able to show, quite surprisingly, that the targeted use of a so-called siRNA tailored specifically to ISG 15 or directed against ISG 15 leads to a significant decrease in the gene activity of ISG 15 and consequently to a significant decrease decreased synthesis or proliferation or replication of hepatitis C virus in the host system leads.
  • the Applicant has succeeded in providing a completely novel and, in addition, low-side-effects therapeutic approach, on the basis of which hepatitis C disease can be effectively treated.
  • the present invention - according to a first aspect of the present invention - is thus the use according to the invention as claimed in claim 1, that is to say in other words the use of an inhibitor and / or repressor of a protein associated with the propagation and / or replication of yeast.
  • Patitis viruses in particular hepatitis C viruses, related nucleic acid molecule, in particular gene and / or its DNA sequence and / or its associated RNA sequence and / or its associated (poly) peptide for the manufacture of a medicament for prophylactic and / or therapeutic treatment of hepatitis, especially hepatitis type C.
  • the central idea of the present invention thus lies in specifically lowering and / or inhibiting, in particular, the gene activity of such a gene, which is associated with the multiplication or synthesis or synthesis of hepatitis C virus.
  • the term "inhibitor” and / or “repressor” used according to the invention refers in particular to a gene activity-reducing and / or suppressing, in particular inhibiting substance.
  • the inhibitor or the repressor thus constitutes a compound which suppresses or at least minimizes the activity of the gene, the pharmacological action of the inhibitor or repressor being at the gene level, ie, for example, by a direct interaction with the gene or its associated Promoters and / or enhancers, as well as at the level of the gene product (s), such as transcription products (eg mRNA) and / or translation products (eg proteins).
  • s such as transcription products (eg mRNA) and / or translation products (eg proteins).
  • the inhibitor or repressor used according to the invention can reduce expression or gene activity, for example, and in a non-limiting manner by direct or indirect interaction with the DNA sequence or DNA and / or its associated RNA sequence or mRNA and / or by direct or indirect influence or interaction with the gene product in the form of the gene or of the DNA Sequence coded (poly) peptide lead.
  • the inhibitor or repressor is in particular a substance with antiviral properties against hepatitis viruses, in particular hepatitis C viruses.
  • the present invention aims significantly at a purposeful suppression of gene expression or gene activity of genes related to the replication or synthesis of hepatitis C viruses, in particular ISGl 5, which in general also as gene knockdown or as gene silencing can be called.
  • the corresponding nucleic acid molecule or the corresponding DNA sequence is likewise included in this regard.
  • the present invention also relates equally, as stated above, to the RNA sequence assigned to the gene, in particular mRNA, which acts as a posttranscriptional product, so to speak, and may be complementary to the co-dogenic strand of the DNA of the gene.
  • the (poly) peptide encoded by the nucleic acid or the gene in particular as previously defined, and thus to a certain extent the translation product, can also be used or used as the target molecule or "target”.
  • nucleic acid molecule is synonymous with the term “polynucleic acid” or “Polynukleinkla- Ie” and may be related to both DNA and RNA.
  • DNA sequence or “RNA sequence” in this context refers not only to the complete, the gene assigned or corresponding DNA, but also to corresponding sections of the gene or not only in particular in the context of transcription synthesized complete RNA, but also on RNA sections or RNA fragments.
  • the gene considered in the context of the use according to the invention is concerned, it is preferably a human gene.
  • the gene considered in the context of the use according to the invention may be an interferon-stimulated gene (ISG).
  • the gene may be a gene stimulated by Type I interferons, preferably ⁇ -interferon and / or ⁇ -interferon. How to- described above, this is in particular a gene which can be induced or activated as a result of an increased interferon concentration, so that its expression is increased, in particular under the influence of interferon, for example caused by an infection with hepatitis C viruses.
  • the Applicant has found, in a completely surprising manner, that a gene responsible for the replication or replication of hepatitis C viruses is formed by the gene ISG 15. This is likewise an interferon-stimulated gene.
  • the gene in question is ISG 15, in particular with the transcript ID (locus) NM 005101 or in particular according to sequence listing I and / or in particular according to SEQUENCE LISTING.
  • ISG 15 which is also referred to as a so-called Homo sapiens ISG 15 ubiquitin-like modifier to a particularly effective in terms of the use of the invention in the context of the therapy of hepatitis type C target or Target, whose inactivation or suppression leads to a significant reduction of hepatitis C viruses in the host system.
  • sequence listing I given above and the previously mentioned SEQUENCE LISTING refer to the DNA sequence to the gene ISG15.
  • the sequence protocol I is synonymous with the corresponding SEQUENCE LISTING or the same content.
  • the essential difference between the sequence protocols is that the sequence protocol I is based on a scientifically standardized specification or presentation, while the SEQUENCE LISTING was created on the basis of a patent-legally standardized specification or representation using the software PatentIN Version 3.3.
  • this is a purely formal representation, but not a difference in content.
  • the inhibitor or the repressor has a gene associated with the gene, in particular with ISGl 5, or with its DNA sequence and / or with its associated RNA sequence and / or with its ( Poly) peptide interacting substance.
  • the term "inhibitor” or “repressor” is to be understood very broadly; in particular, it may be a substance which directly and / or indirectly, for example via metabolic cascades or signal transduction, with the corresponding target or Target, in particular with ISGl 5, interacts and thereby reduces or inhibits its gene activity, in particular inhibited.
  • the influencing of the gene, in particular of ISGl 5, or of the gene or ISG 15 associated products can also be caused in the form of precursor substances, for example, only in the cell or Host system is converted to the actually interacting substance, for example via metabolism-specific processes.
  • the direct and / or indirect interaction of the substance with the target structure, in particular with the gene, such as ISG 15, can be realized on many levels:
  • the interacting substance or the inhibitor and / or repressor with the promoter and / or enhancer of the gene, in particular of ISG 15, interact and / or interact such that the binding of particular endogenous transcription factors, in particular activators to which promoter and / or enhancer is prevented or at least inhibited.
  • the substance or the inhibitor and / or repressor with a particular endogenous transcription factor, in particular activator interacts such that the binding of the transcription factor, in particular activator, to the promoter and / or enhancer of the gene, in particular of ISGl 5, prevented or at least inhibited.
  • the substance or the inhibitor and / or repressor with a particular endogenous transcription factor, in particular activator interacts such that the binding of the transcription factor, in particular activator, to the promoter and / or enhancer of the gene , in particular of ISG 15, is prevented or at least inhibited.
  • the inhibitor and / or repressor may be used with, in particular, endogenous, mediator-regulated mediators and / or factors, in particular ISG15-regulated mediators and / or factors, and / or with, in particular, endogenous gene regulating mediators and / or factors.
  • ISG 15-regulating mediators and / or factors interact in particular such that the activity of the gene, in particular of ISGl 5, and / or its DNA sequence and / or its associated RNA sequence and / or its associated (Poly -) Peptides reduced and / or prevented, in particular inhibited is.
  • the substance or the inhibitor and / or repressor reacts with the endogenous or endogenous transcription activators themselves and thereby causes an inactivation of the transcription factor or activator per se, in order in this way to inhibit the gene activity, in particular of ISG 15, to reduce or to prevent, in particular to inhibit.
  • the substance used in the context of the use according to the invention in the form of an inhibitor and / or repressor may be, on the one hand, a substance which has been found on the basis of the method for the identification of such substances which is described below.
  • the inhibitor and / or repressor on the other hand may be an RNA sequence which interacts preferentially with the RNA sequence, in particular ISGl 5, associated with the gene, in particular mRNA sequence.
  • the inhibitor or repressor should be an RNA or RNA sequence which is in particular complementary to the RNA or mRNA or RNA sequence or mRNA sequence assigned to the gene.
  • the inhibitor and / or repressor prefferably have an RNA sequence in the form of an oligomer, in particular of 15 to 20 bp (base pairs), preferably 18 to 25 bp, preferably 21 to 23 bp, is.
  • the RNA sequence which interacts with the RNA sequence assigned to the gene, in particular ISG1 is preferably a single-stranded RNA, and in this respect according to a particularly preferred embodiment is a so-called antisense strain, in particular with respect to the RNA associated with the gene, in particular mRNA.
  • RNA sequence interacting with the RNA sequence, which interacts with the gene, in particular ISGl 5 to be a double-stranded RNA which, in particular in the context of further modifications or metabolic processes, becomes a Single-stranded RNA, in particular in the form of an antisense strand, as previously defined, converted or modified.
  • the inhibitor and / or repressor described above is an siRNA (small interfering RNA), in particular wherein the siRNA is directed against ISG15, in particular against ISG15-specific mRNA or mRNA associated with ISG15 is.
  • the siRNA should be designed such that an interaction with the mRNA, in particular as defined above, is possible and consequently leads to the deactivation or to the degradation of the corresponding RNA.
  • the siRNA may be complementary to the mRNA or to sections of the mRNA.
  • the Applicant has found, in a completely surprising manner, that such an siRNA leads to particularly good results with regard to a reduction or inhibition of the gene activity of the gene associated with the multiplication or replication of hepatitis C viruses, in particular ISGl 5, leads.
  • siRNA used in the context of the use according to the invention is, according to a very particularly preferred embodiment, an siRNA against ISGl 5, which is available from Qiagen, Hilden, Germany under the product number or order number SI00072387 (human) or SIO 1007531 ( murine) can be obtained.
  • the mode of action of the specific ISG15-specific siRNA can be understood in such a way that initially an siRNA / protein complex is formed in the cell or host system, which is also called an RNA-induced silencing complex (RISC), in which a protein complex binds the antisense strand of the siRNA and cuts the complementary mRNA.
  • RISC RNA-induced silencing complex
  • the RISC complex has RNA helicase and RNA nuclease activities.
  • the inhibitor and / or repressor is an inhibitor or repressor, in particular the antisense strand of an siRNA.
  • a central concept of the present invention according to this aspect of the invention using a siRNA is to be seen in that in particular synthetically produced siRNA with specificity to mRNA as a transcription product in particular of ISG 15 is introduced into the cell or host system, which leads to a degradation of the mRNA of the target gene, namely in particular ISG 15 leads, which in turn leads to a reduction of the gene products and thus to a genes ilencing or gene knockout.
  • the principle underlying the invention according to this embodiment may also be referred to as RNA interference, based on which the expression of certain (target) genes, in this case in particular ISG 15, is reduced. This occurs in particular through the interaction of siRNA directed specifically against a gene with the mRNA of the gene with the consequence of the degradation of the mRNA, which corresponds to a (gene) inactivation.
  • the introduction or transfecting can also be carried out by means of a so-called polyethylenimine complexation (PEI complexing).
  • PEI complexing polyethylenimine complexation
  • the inhibitor or repressor used according to the use of the present invention should be administered when administered in pharmaceutically effective amounts.
  • the inhibitor or the repressor should be administered systemically, for example intravenously.
  • the measures relating to this are also known to the person skilled in the art, so that no further explanation is required in this respect.
  • the inhibitor or repressor in particular as described above, is administered together with at least one interferon, in particular ⁇ -interferon, preferably pegylated ⁇ -interferon.
  • interferon in particular ⁇ -interferon, preferably pegylated ⁇ -interferon.
  • a combination of the previously described ISGl 5 -specific siRNA or ISG 15-directed siRNA with an interferon is of particular advantage. Because the Applicant could - as stated below with reference to the embodiments - in a completely surprising manner, a synergistic effect in the context of the combination described above. on the one hand and interferon on the other hand with regard to the suppression or reduction of the multiplication or replication of hepatitis C viruses in the affected host systems or cell systems. Likewise, it is possible for the inhibitor and / or repressor to be administered together and / or in combination with at least one substance having antiviral properties to hepatitis viruses, in particular hepatitis C viruses.
  • a new way of treating a hepatitis C infection or disease is provided, which is based on a completely new approach, namely the targeted inactivation of one in connection with the multiplication or Replication of hepatitis C viruses, especially ISGl 5.
  • Another object of the present invention - according to one aspect of the present invention - is the use according to the invention according to claim 15, d. H.
  • Another object of the present invention - according to a third aspect of the present invention - is the use according to claim 17, ie in other words a use of at least one interferon-stimulated nucleic acid molecule, in particular gene, preferably ISGl 5, and / or its DNA sequence and / or its associated RNA sequence and / or at least one of nucleic acid-encoded (poly) peptide for finding and / or providing a medicament for the prophylactic and / or curative treatment of hepatitis, in particular hepatitis type C, and / or for predicting individual drug effects and / or drug side effects.
  • interferon-stimulated nucleic acid molecule in particular gene, preferably ISGl 5, and / or its DNA sequence and / or its associated RNA sequence and / or at least one of nucleic acid-encoded (poly) peptide for finding and / or providing a medicament for the prophylactic and / or curative treatment of hepatitis, in
  • the gene associated with the multiplication or replication of hepatitis C viruses can be regarded as a starting material for the discovery or provision of medicaments with respect to a hepatitis C infection or disease are used.
  • the medicaments can be such that, as stated above, they interact directly or indirectly with the previously defined gene or the RNA associated therewith, in particular mRNA, and / or the corresponding (poly) peptide in order in this way in particular via a reduction or inhibition, in particular inhibition, of the gene activity to reduce or prevent the multiplication or replication of hepatitis C viruses and thus to alleviate or cure the hepatitis C disease.
  • These may thus be substances which have a gene regulatory effect.
  • Another object of the present invention - according to a fourth aspect of the present invention - is the inventive method according to claim 18, ie in other words a method for identifying an inhibitor and / or repressor of an interferon-stimulated nucleic acid molecule, in particular gene, preferably of ISG 15, and / or its DNA sequence and / or its associated RNA sequence, the method comprising the following steps: (A) contacting the nucleic acid molecule with at least one test substance under conditions which allow an interaction, in particular binding, of the test substance (s) to the nucleic acid molecule; and (b) detecting and / or analyzing whether the test substance (s) limit or prevent the gene activity and / or expression of the nucleic acid molecule and / or whether the test substance (s) increase the proliferation and / or replication of hepatitis viruses, in particular Restrict or prevent hepatitis C viruses.
  • the method according to the invention can be carried out, for example, in vitro, to a certain extent investigating an interaction of the test substance with the nucleic acid molecule or the gene and, in the presence of interaction, concluding that the gene activity as a result of this interaction is reduced.
  • the method according to the invention can equally be carried out in a corresponding host system, wherein the host should be a carrier of the gene associated with the multiplication or replication of hepatitis C viruses, in particular ISG 15, and advantageously also via a corresponding expression system features.
  • the detection of the interaction or interaction or the detection of the influence of the gene activity can be carried out by methods familiar to the person skilled in the art.
  • the present invention relates to a method according to claim 19, d. H.
  • a method for identifying an inhibitor and / or repressor of a (poly) peptide encoded by an interferon-stimulated nucleic acid molecule, in particular gene, preferably ISG 15, and / or its DNA sequence and / or its associated RNA sequence wherein the method comprises the following steps:
  • the inventive method according to this aspect thus focuses on influencing the gene product, in particular in the form of a protein or (poly) peptide.
  • the method can be carried out in a manner known to those skilled in the art both in vitro and in vivo in a host system, wherein in the latter case the host should preferably carry the nucleic acid encoding the (poly) peptide to be examined.
  • an interaction can also be carried out in vitro on isolated (poly) peptides.
  • the present invention likewise relates to a method for identifying a mediator and / or factor which is regulated by an interferon-stimulated nucleic acid molecule, in particular gene, preferably ISG 15, or which regulates an interferon-stimulated nucleic acid molecule, in particular gene, preferably ISGl 5 , interacting substance, the method comprising the following steps:
  • test substance (s) influence the activity of the mediator and / or factor and / or whether the test substance (s) is responsible for the multiplication and / or replication of hepatitis viruses, in particular hepatitis C; -Viren, restrict or prevent.
  • the interacting substance is preferably an inhibitor and / or repressor of the mediator and / or factor, provided that this is associated with an activation of the interferon-stimulated nucleic acid molecule, in particular gene, preferably ISG 15.
  • the mediator and / or factor itself is in connection with an inactivation or repression of the interferon-stimulated nucleic acid molecule, in particular gene, preferably ISG 15, this is the case the interacting substance with respect to the mediator and / or factor to an activator - with respect to the interferon-stimulated nucleic acid, in particular gene, preferably ISGl 5, based on the above definition in such a substance, however, is equally an inhibitor or Repressor, as in this case ultimately results in inactivation or repression of the interferon-stimulated nucleic acid molecule, in particular gene, preferably ISG 15 results.
  • inventive methods according to the fourth and fifth aspects of the present invention can be carried out such that several test substances are used and the following steps are carried out:
  • RNA in particular mRNA
  • hepatitis C viruses restrict or not prevent and / or which do not interact with the mediator and / or factor in the further test procedure are no longer considered;
  • step (c) repeating step (b) until a single test substance is identified which comprises reducing or inhibiting gene activity and / or expression of the nucleic acid molecule and / or reducing or inhibiting the activity of the (poly) peptide and / or the reduction or elimination of multiplication and / or Replication of hepatitis viruses, in particular hepatitis C viruses, and / or to which the interaction with the mediator and / or factor can be assigned.
  • the method according to the fourth and fifth aspect of the present invention be carried out such that the test substance (s), the nucleic acid molecule (DNA or RNA, in particular mRNA) and / or the (poly) peptide to a Readout system (readout system) and / or that a readout system is added to the test batch and / or that the readout system after binding of the test substance (s) with the nucleic acid molecule and / or the (poly) peptide is a detectable signal supplies.
  • a Readout system readout system
  • a readout system is added to the test batch and / or that the readout system after binding of the test substance (s) with the nucleic acid molecule and / or the (poly) peptide is a detectable signal supplies.
  • test substances may be low-molecular substances, peptides, aptamers, antibodies, DNA, RNA, in particular siRNA and / or fragments or derivatives thereof.
  • the aforementioned methods can be carried out, for example, in a host or host system, wherein the host or the host system preferably comprises the previously defined genes and has a corresponding expression system.
  • host or the host system preferably comprises the previously defined genes and has a corresponding expression system.
  • Such hosts are familiar to those skilled in the art or the skilled person is always able to select specific host systems in the context of the present invention, so that there is no further explanation in this regard.
  • the inventive methods can likewise be carried out in the form of high-throughput methods and / or computer-assisted.
  • test substance (c) modifying the test substance such that its binding specificity or binding affinity or binding avidity is increased for the nucleic acid molecule or the (poly) peptide.
  • the binding site in step (a) can be determined by site-directed mutagenesis, the procedures of which are known per se to those skilled in the art.
  • Yet another subject of the present invention - according to one aspect of the present invention - is the inventive method according to claim 28, d. H. in other words, a method for modifying a test substance which is identified or improved according to the methods according to the fourth and / or fifth and / or sixth aspect of the present invention, wherein the test substance is further modified as a lead structure
  • the identified, improved or modified test substance in particular the inhibitor and / or repressor of the aforementioned gene, to be further pharmacologically improved by peptide mimetics.
  • Yet another subject of the present invention - according to an aspect of the present invention - is the method according to the invention according to claim 30, d.
  • a method for the identification and / or determination of at least one nucleic acid molecule associated with the replication of hepatitis viruses, in particular hepatitis C viruses, in particular gene, preferably human and / or interferon-stimulated gene, the method comprising the following Steps includes:
  • the method may comprise, subsequent to step (c), the following step (d):
  • step (d) assignment of the nucleic acid molecule identified in step (c), in particular gene, as a nucleic acid molecule, in particular gene, associated with the multiplication and / or replication of hepatitis viruses, in particular hepatitis C viruses.
  • the inventive method according to this aspect of the present invention can be used, for example, by differential expression using so-called DNA chips.
  • DNA chips DNA chips
  • Analysis method the expression level in subjects with hepatitis C infection compared to subjects without hepatitis C infection for certain genes is determined and a gene, in particular an interferon-stimulated gene, with an increased expression level, the properties of a in connection with the multiplication or .
  • Replication of hepatitis C virus-related gene, in particular the multiplication or replication of hepatitis C virus-promoting gene can be awarded.
  • genes in particular interferon-stimulated genes, which are associated with the multiplication or replication of hepatitis C viruses in connection with a hepatitis C infection or hepatitis C disease.
  • the present invention according to a ninth aspect of the present invention relates to a pharmaceutical composition according to claim 32, in other words, a pharmaceutical composition, preferably for the prophylactic or therapeutic treatment of hepatitis C diseases, containing in effective, in particular pharmaceutically active Quantities of at least one pharmacokinetic gically active substance, in particular an inhibitor and / or repressor, for a nucleic acid molecule in connection with the multiplication and / or replication of hepatitis viruses, in particular hepatitis C viruses, in particular gene, preferably ISGl 5, and / or for the latter DNA sequence and / or its associated RNA sequence, in particular for its associated mRNA sequence, and / or for its associated (poly) peptide, wherein the substance according to the method according to the fourth and / or fifth and / or sixth Aspect of the present invention is available.
  • a pharmaceutical composition preferably for the prophylactic or therapeutic treatment of hepatitis C diseases, containing in effective, in particular pharmaceutically active Quantities of at
  • Yet another object of the present invention - according to a tenth aspect of the present invention - is also a pharmaceutical composition according to claim 33, in other words a pharmaceutical composition, preferably for the prophylactic or therapeutic treatment of hepatitis C diseases, containing in effective, in particular pharmaceutically effective amounts of at least one siRNA, wherein the siRNA regulates, in particular reduces or at least inhibits, the gene activity and / or gene expression of at least one interferon-stimulated gene, in particular of ISG 15.
  • the siRNA used in the context of the pharmaceutical composition according to the invention is an siRNA against ISG 15, which can be obtained from Qiagen, Hilden, Germany under the product number or order number SI00072387.
  • the cells were overlaid with 500 ⁇ l of trizole.
  • the adherent cells were detached from the bottom with a plastic scraper and transferred to a reaction vessel.
  • 0.1 ml of chloroform / 1 ml of trizole was added and mixed by shaking. Centrifugation at 12,000 g and at 2 to 8 0 C for 15 min resulted in the phase separation of the phenol / chloroform mixture.
  • the aqueous phase was removed and the dissolved RNA precipitated by 0.5 ml of isopropanol / 1 ml of trizole.
  • the pellet was then washed with 75% ethanol, dried under vacuum and finally dissolved in RNase-free water. Subsequently, the RNA was assessed using the "RNeasy Mini" kit (Qiagen, Hilden, Germany) purified according to the manufacturer's instructions and stored until further analysis at -20 0 C.
  • Quantitative real-time PCR The reverse transcription of RNA, followed by a polymerase chain reaction (RT-PCR), is a sensitive method for the quantification of specific mRNAs.
  • RT-PCR polymerase chain reaction
  • the use of specific primers first transcribes the mRNA of the gene in question into complementary DNA (cDNA), which in turn is the template or the template for the following PCR delivers.
  • cDNA complementary DNA
  • the LightCycler Rotor-Gene 2000 was used by Corbett (Mortlake, Australia).
  • the LightCycler uses a fluorimeter component to detect the fluorescence of the fluorophore after binding to double-stranded DNA.
  • QIAGEN's QuantiTect SYBR Green RT-PCR kit was used, and a 25 ⁇ l mixture was pipetted together as follows: 5.25 ⁇ l H 2 O (RNase-free), 12.5 ⁇ l SYBR Green RT-PCR Master Mix, 0 , 25 ⁇ l of QuantiTect RT Mix, 2.5 ⁇ l of each primer (0.5 mM) and 2 ⁇ l total RNA (100 ng to 200 ng).
  • the LightCycler program used started with a 30 minute RT step at 55 0 C, followed by a 15 minute heat inactivation of the RT-polymerase of a conventional terminal with the PCR schemes, ie 5 s denaturation per cycle at 95 ° C., 10 s annealing temperature (55 ° C.) and 30 s elongation at 72 ° C.
  • Product formation was determined by fluorescence increase after each replication cycle. After an average of 40 replication cycles, the melting curves of the formed products were recorded to check the specificity of the PCR reaction. Due to the melting behavior of DNA, the fluorescence decreases with increasing temperature. The maximum fluorescence change per temperature increase gives a maximum in the melting curve, which is characteristic of each PCR product.
  • the copy numbers of the measured genes calculated by the LightCycler were compared with the housekeeping gene or ß-actin gene and analyzed.
  • the expression of ISGl 5 was switched off at the cell culture level in the HCV replicon system in order to investigate the direct effect on HCV replication.
  • the suppression of gene expression takes place by means of siRNA via a cellular mechanism of processing.
  • the Dicer enzyme complex cleaves cell-type dsRNA into 21 to 23 bp oligomers, the so-called small interfering RNAs (siRNA).
  • the siRNA can form the RNA-induced silencing complex (RISC) with a protein complex; this binds the antisense strand of the siRNA and cuts the complementary mRNA.
  • RISC RNA-induced silencing complex
  • the degradation of the mRNA leads to a reduction in the translation of these mRNAs and thus to gene silencing at the post-transcriptional level.
  • siRNAs 12.5 ng were first dissolved in 3 ⁇ l Susspensionspuffer and pre-pipetted into the wells of the perforated plate in a reverse transfection.
  • 2.5 ng of the siRNAs were used for simultaneously switching off other genes (not shown graphically).
  • a noncodogenic siRNA was inserted. puts.
  • 0.75 ⁇ l of the transfection reagent Hi-PerFect TM 0.75 ⁇ l of the transfection reagent Hi-PerFect TM (Qiagen, Hilden, Germany) was transferred to 24.25 ⁇ l serum-free culture medium per batch, mixed and added to the siRNAs that were initially introduced. The transfection mixture was then incubated for 10 min at room temperature.
  • Figure IA the HCV copy number is normalized and aligned against the untreated control (MHI) and in Figure IB the HCV / ISG15 copy number is normalized and aligned against the siNC control.
  • Figures IA and IB show the significant decrease in HCV copy number when treated with ISG15-specific siRNA (siISG15). Consequently, the replication under the influence of SÜSG15 is also reduced.
  • the murine and human MHL conl-HCV replicon cells were seeded in 6-well plates and grown to a confluency of 30 to 40% at 37 0 C and a CO incubated for 2 -Luftgehalt of 5%.
  • the cells were washed and taken up in culture medium as well as with IFN- ⁇ -added medium and incubated for an additional 64th Subsequently, the cells were lysed and a protein extraction was performed.
  • the viral protein NS5A and the housekeeping gene ß-actin were detected by means of Western blot.
  • Figures 2A and 2B show in a Western blot that treatment with anti-ISGl 5 siRNA in both murine and human HCV replicon systems results in significant suppression of the HCV NS5A protein. Further, Figures 2A and 2B show that there is synergism with IFN- ⁇ since the additional administration of IFN- ⁇ (see Figures 2A and 2B: + IFN- ⁇ ) results in a significant reduction in NS5A-. Synthesis compared to the non-IFN- ⁇ treated approach (see Figures 2A and 2B: IFN- ⁇ ).
  • Fig. IA shows the inhibition of HCV replication and the lack of induction of resistant mutants when treated with ISG15-specific siRNA (siISG15) versus treatment with siNC, where the HCV copy number is normalized and balanced against the untreated control (MHI).
  • Figure IB shows the inhibition of HCV replication and the lack of induction of resistant mutants when treated with ISG15-specific siRNA (siISG15) versus siNC treatment
  • HCV / ISG 15 copy number is normalized and balanced against the siNC control.
  • Fig. 2A shows by means of a Western blot the results of a treatment performed on human conl-HCV replicon cells with gegen
  • FIG. 2B shows the results of a mouse on a western blot
  • ISG 15 directed siRNA without additional application of IFN- ⁇ on the one hand (- IFN- ⁇ ) and with additional application of IFN- ⁇ on the other hand (+ IFN- ⁇ ).
  • MHl untreated control
  • NC non-silencing control
  • ISG 15 ISG15-specific si-RNA

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Genetics & Genomics (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Molecular Biology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • Biotechnology (AREA)
  • General Health & Medical Sciences (AREA)
  • Biomedical Technology (AREA)
  • Communicable Diseases (AREA)
  • Virology (AREA)
  • Biophysics (AREA)
  • Biochemistry (AREA)
  • Microbiology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Immunology (AREA)
  • Physics & Mathematics (AREA)
  • Medicinal Chemistry (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Oncology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Animal Behavior & Ethology (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Analytical Chemistry (AREA)
  • Plant Pathology (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

L'invention concerne le domaine du traitement des infections de type hépatite ou des maladies relevant de l'hépatite, en particulier de l'hépatite type C. En particulier, l'invention a pour objet l'utilisation d'un inhibiteur et/ou d'un répresseur d'une molécule d'acide nucléique, en particulier d'un gène, en rapport avec la prolifération et/ou la réplication de virus de l'hépatite, en particulier de virus de l'hépatite C, pour la production d'un médicament destiné au traitement prophylactique et/ou curatif de l'hépatite, en particulier de l'hépatite C. L'invention concerne également une composition pharmaceutique, de préférence pour le traitement prophylactique ou thérapeutique de maladies relevant de l'hépatite C, contenant un répresseur et/ou un inhibiteur.
EP09745480A 2008-05-16 2009-03-12 Nouveaux agents thérapeutiques pour le traitement de l'hépatite Withdrawn EP2285962A2 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
DE102008024010 2008-05-16
DE102008029669A DE102008029669A1 (de) 2008-05-16 2008-06-24 Neue Therapeutika für die Hepatitis-Therapie
PCT/EP2009/001780 WO2009138146A2 (fr) 2008-05-16 2009-03-12 Nouveaux agents thérapeutiques pour le traitement de l'hépatite

Publications (1)

Publication Number Publication Date
EP2285962A2 true EP2285962A2 (fr) 2011-02-23

Family

ID=41180545

Family Applications (1)

Application Number Title Priority Date Filing Date
EP09745480A Withdrawn EP2285962A2 (fr) 2008-05-16 2009-03-12 Nouveaux agents thérapeutiques pour le traitement de l'hépatite

Country Status (5)

Country Link
US (1) US20110076251A1 (fr)
EP (1) EP2285962A2 (fr)
CN (1) CN102099473A (fr)
DE (1) DE102008029669A1 (fr)
WO (1) WO2009138146A2 (fr)

Families Citing this family (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DK2583677T1 (da) 2011-10-21 2015-01-19 Abbvie Inc Fremgangsmåder til behandling af HCV omfattende mindst to direktevirkende antivirale midler, ribavirin, men ikke inteferon
US8466159B2 (en) 2011-10-21 2013-06-18 Abbvie Inc. Methods for treating HCV
DK2583680T1 (da) 2011-10-21 2015-01-19 Abbvie Inc Mono (PSI-7977) eller kombinationsbehandling af DAA til anvendelse ved behandling af HCV
US8492386B2 (en) 2011-10-21 2013-07-23 Abbvie Inc. Methods for treating HCV
EP3448392A4 (fr) 2016-04-28 2020-01-15 Emory University Compositions thérapeutiques à base de nucléotides et nucléosides contenant un alcyne et utilisations associées
CN110241198A (zh) * 2019-05-30 2019-09-17 成都吉诺迈尔生物科技有限公司 一种表征hHRD同源重组缺陷的基因组重组指纹及其鉴定方法

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7608600B2 (en) * 2002-06-28 2009-10-27 Idenix Pharmaceuticals, Inc. Modified 2′ and 3′-nucleoside prodrugs for treating Flaviviridae infections
WO2006006948A2 (fr) * 2002-11-14 2006-01-19 Dharmacon, Inc. Methodes et compositions permettant de selectionner des arnsi presentant une fonctionnalite amelioree
WO2004065577A2 (fr) * 2003-01-14 2004-08-05 Bristol-Myers Squibb Company Polynucleotides et polypeptides associes a la voie nf-$g(k)b
WO2004069860A2 (fr) * 2003-02-03 2004-08-19 The Scripps Research Institute Proteines conjuguees a isg15
US20100248230A1 (en) * 2007-04-14 2010-09-30 Schlaak Joerg Friedrich Novel medical diagnostic method and therapy in the context of interfreon-stimulated genes that induce depression

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO2009138146A2 *

Also Published As

Publication number Publication date
US20110076251A1 (en) 2011-03-31
DE102008029669A1 (de) 2009-11-19
CN102099473A (zh) 2011-06-15
WO2009138146A3 (fr) 2010-04-08
WO2009138146A2 (fr) 2009-11-19

Similar Documents

Publication Publication Date Title
Chen et al. LncRNA GAS5 regulates ischemic stroke as a competing endogenous RNA for miR-137 to regulate the Notch1 signaling pathway
EP2684956B1 (fr) Procédé pour stimuler l'angiogenèse, la vascularisation ou la réparation vasculaire ou pour empêcher l'angiogenèse tumorale
DE10163098B4 (de) Verfahren zur Hemmung der Replikation von Viren
EP2285962A2 (fr) Nouveaux agents thérapeutiques pour le traitement de l'hépatite
WO2013127782A2 (fr) Utilisation de micro-arn ou de gènes comme marqueurs pour l'identification, le diagnostic et le traitement de formes individuelles de cardiomyopathies non ischémiques ou de maladies de surcharge du cœur
WO2003035876A1 (fr) Utilisation d'un acide ribonucleique a double brin pour traiter une infection a virus a arn a simple brin positif
WO2003035083A1 (fr) Medicament destine a traiter une fibrose par interference d'arn
KR102105016B1 (ko) miR-485-3p를 이용한 알츠하이머병 진단 방법
Zhang et al. Tanshinone IIA ameliorates dextran sulfate sodium-induced inflammatory bowel disease via the pregnane X receptor
Liu et al. Silencing microRNA‑29b‑3p expression protects human trabecular meshwork cells against oxidative injury via upregulation of RNF138 to activate the ERK pathway
WO2021107005A1 (fr) Agent prophylactique ou thérapeutique pour des maladies associées au cytomégalovirus
Chen et al. Anti‑inflammatory effects of miR‑150 are associated with the downregulation of STAT1 in macrophages following lipopolysaccharide treatment
CN107137710B (zh) Kindlin-2蛋白作为靶点在制备治疗肾炎的药物中的应用
AU2009289239B2 (en) Treatment of scleroderma
US20200016189A1 (en) Method for treating schizophrenia
Stonedahl et al. Treatment with granulocyte-macrophage colony-stimulating factor reduces viral titers in the brains of west nile virus-infected mice and improves survival
Li et al. Remote ischemic conditioning alleviates chronic cerebral hypoperfusion-induced cognitive decline and synaptic dysfunction via the miR-218a-5p/SHANK2 pathway
AT509045A4 (de) Verbindungen zur behandlung von asthma bronchiale
Makowska et al. Sequential induction of type I and II interferons mediates a long-lasting gene induction in the liver in response to a novel toll-like receptor 9 agonist
DE102008017701A1 (de) Neue medizinische Diagnostik und Therapie im Zusammenhang mit interferonstimulierten depressionsvermittelnden Genen
KR20200043322A (ko) miR-485-3p를 이용한 알츠하이머병 진단 방법
US9951336B2 (en) Composition for reducing senescence of cell or subject comprising smurf2 inhibitor and use thereof
CN116585326B (zh) 科罗索酸在制备抑制流行性乙型脑炎病毒药物中的用途
KR102645546B1 (ko) 간성뇌증에 의해 유발되는 신경학적 또는 정신학적 장애 관련 질환 진단용 마커 및 이를 이용한 간성뇌증에 의해 유발되는 신경학적 또는 정신학적 장애 관련 질환 진단에 필요한 정보를 제공하는 방법
US12084718B2 (en) Method for diagnosis of Alzheimer's disease using microrna

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 20101108

AK Designated contracting states

Kind code of ref document: A2

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LI LT LU LV MC MK MT NL NO PL PT RO SE SI SK TR

AX Request for extension of the european patent

Extension state: AL BA RS

17Q First examination report despatched

Effective date: 20120918

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN

18D Application deemed to be withdrawn

Effective date: 20140408