KR20220011349A - Liver cancer specific biomarkers and use thereof - Google Patents

Abstract

The present invention relates to a use of using, as biomarkers for the detection and diagnosis of hepatocellular carcinoma, genes of which the expression changes specifically to hepatocellular carcinoma, and treating liver cancer by using the same as targets. In the present invention, it has been discovered that the expression of SMARCA4 is increased in HCC and SMARCA4 directly increases the expression of IRAK1, and it has been ascertained that Gankyrin and AKR1B10, which are oncogenic proteins, are induced through the transcriptional activation of IRAK1, and thus SMARCA4-IRAK1-Gankyrin and AKR1B10 can be used as markers specific for liver cancer and can be effectively used as targets for liver cancer treatment.

Description

Liver cancer-specific biomarkers and their uses {LIVER CANCER SPECIFIC BIOMARKERS AND USE THEREOF}

The present invention relates to a liver cancer-specific biomarker and its use, and more particularly, using genes whose expression is specifically changed in hepatocellular carcinoma as biomarkers for hepatocellular carcinoma detection and diagnosis, and to treat liver cancer by targeting them It's about use.

Cancer is a representative disease that threatens human health and is the most representative cause of death as a single disease in industrialized countries. Although the cause of cancer is still unknown, it is considered that complex factors of genetic factors, which are internal factors, and carcinogenic chemicals acting as factors that cause cancer, which are external factors, continuous inflammation and damage, and cancer-causing virus infection are acting. . However, cancer is not so desperate that it can be concluded that it is an incurable disease, and it can be cured by early diagnosis and active treatment. Therefore, early detection, early diagnosis, and treatment are crucially important to increase the effectiveness of cancer treatment, and even in advanced cancer, if various and active methods are used, a cure or life can be prolonged and painful symptoms can be improved.

Among cancers, liver cancer is known as one of the deadliest cancers in the world, and its mortality rate is the third most aggressive cancer (Ahn J, Flamm SL Hepatocellular carcinoma Dis Mon 2004;50:556-573). It is possible only in % to 25% of patients, and most liver cancer patients die within a relatively short period of time due to locally advanced or metastatic diseases (Roberts LR, Gores GJ Hepatocellular carcinoma: molecular pathways and new therapeutic targets Semin Liver Dis 2005; 25:212-225) Hepatitis B virus, hepatitis C virus, and aflatoxin B1 are well known as major causes of liver cancer. However, the overall survival rate of liver cancer patients has not increased significantly over the past 20 years, and the mechanism of development and progression of liver cancer is still not well known (Bruix J, et al Focus on hepatocellular carcinoma Cancer Cell 2004; 5:215-219) So far, molecular targeted therapy has been shown to be effective for the treatment of mature liver cancer (Shen YC, Hsu C, Cheng AL Molecular targeted therapy for advanced hepatocellular carcinoma: current status and future perspectives J Gastroenterol;45:794-807), it is unclear how these genetic alterations lead to the clinical features observed in individual HCC patients. Liver cancer can be broadly divided into primary liver cancer (hepatocellular carcinoma) arising from hepatocellular carcinoma and metastatic liver cancer in which cancers from other tissues have metastasized to the liver. More than 90% of liver cancers are primary liver cancer.

Hepatocellular carcinoma (HCC) is the fifth most common tumor in the world, with 500,000 deaths annually (Okuda 2000). Survival rates in HCC patients have not improved over the past 20 years and have an incidence nearly equal to mortality (Marrero, Fontana et al 2005). Chronic hepatitis caused by infection with hepatitis B virus or hepatitis C virus and exposure to carcinogens such as aflatoxin B1 are known as major risk factors for HCC (Thorgeirsson and Grisham 2002), and cell cycle mechanisms There is also a report that changes in cell cycle regulators that progress to the G1 phase are involved in the formation of hepatocarcinoma (Hui et al, Hepatogasteroenterology 45:1635-1642, 1998), but the intracellular molecular mechanisms related to the onset and progression of liver cancer are not. It is still not clearly clarified. According to prior studies, protooncogenes such as various growth factor genes are mutated into oncogenes for various reasons and overexpress or overactive, or tumors such as Rb protein or p53 protein It has been reported that when a tumor suppressor gene is mutated by various causes and underexpressed or loses its function, it causes the onset and progression of various cancers including liver cancer. In addition, it has been reported that DNA mutations and genetic alteration of gene expression are confirmed in liver cancer patient tissues (Park et al, Cancer Res 59:307-310, 1999; Bjersing et al, J Intern Med 234:339). -340,1993; Tsopanomichalou et al, Liver 19:305-311, 1999; Kusano et al, Hepatology 29:1858-1862, 1999; Keck et al, Cancer Genet Cytogenet 111:37-44, 1999). Recently, it is recognized that the onset and progression of most cancers, including liver cancer, is caused by the complex interaction of various genes related to the cell cycle, signal transduction, etc., rather than by a few specific genes. Therefore, individual genes or proteins Rather than focusing only on the expression or function of

It is an object of the present invention to provide a biomarker for diagnosing liver cancer.

In addition, an object of the present invention is to provide a composition for diagnosing liver cancer

Another object of the present invention is to provide a liver cancer diagnostic kit.

Another object of the present invention is to provide a pharmaceutical composition for preventing or treating liver cancer.

Another object of the present invention is to provide a method for providing information necessary for diagnosing liver cancer.

Another object of the present invention is to provide a method for providing information for predicting the prognosis of liver cancer.

In addition, it is an object of the present invention to provide a method for predicting liver cancer treatment responsiveness of a SMARCA4 inhibitor.

In order to achieve the above object, the present invention provides a biomarker for diagnosing liver cancer comprising one or more genes selected from the group consisting of SMARCA4, SMARCC1, SMARCA2 and IRAK1 or proteins expressed from the genes.

In addition, the present invention provides a composition for diagnosing liver cancer comprising an agent for measuring the expression level of one or more biomarker genes selected from the group consisting of SMARCA4, SMARCC1, SMARCA2 and IRAK1 at the mRNA or protein level.

In addition, the present invention provides a liver cancer diagnostic kit comprising the composition.

In addition, the present invention provides a pharmaceutical composition for preventing or treating liver cancer comprising a SMARCA4 inhibitor as an active ingredient.

In addition, the present invention provides a method for providing information necessary for diagnosing liver cancer.

In addition, the present invention provides a method of providing information for predicting the prognosis of liver cancer.

In addition, the present invention provides a method for predicting liver cancer treatment responsiveness of a SMARCA4 inhibitor.

According to the present invention, the expression of SMARCA4 was increased in HCC, it was found that SMARCA4 directly increased the expression of IRAK1, and by confirming that the oncoproteins Gankyrin and AKR1B10 were induced through transcriptional activation of IRAK1, SMARCA4-IRAK1- Gankyrin and AKR1B10 can be used as specific markers for liver cancer, and they can be usefully used as targets for liver cancer treatment.

1 is a diagram confirming the genetic variation and abnormal expression of the SWI / SNF subunit gene in HCC:
A: mutation rate of SWI/SNF complex subunit gene in HCC;
B: Differential gene expression of SMARCA4 , SMARCC1 and SMARCA2 in HCC patients compared to healthy normal patients (NC) in TCGA_LIHC, ICGC_LIRI and Catholic_mLIHC (GSE114654) data set;
C: Differential gene expression of SMARCA4 , SMARCC1 and SMARCA2 in tissue pairs of HCC tissues and corresponding noncancerous liver biopsies;
D: Expression of SMARCA4 , SMARCC1 and SMARCA2 in pairs of HCC tissues analyzed by qRT-PCR and corresponding tissues of HCC from noncancerous liver biopsies; and
E: Western blot analysis results of SMARCA4, SMARCC1 and SMARCA2 in 10 pairs of selected HCC subsets.
2 is a diagram analyzing the tumorigenicity of SMARCA4, SMARCC1 and SMARCA2 in HCC:
A: Cell viability of si-SMARCA4-treated HCC;
B: cell proliferation rate of si-SMARCA4-treated HCC;
C: cell viability of si-SMARCC1-treated HCC;
D: cell proliferation rate of HCC treated with si-SMARCC1; and
E: Cell viability of SMARCA4 high-expressing cells and SMARCA4 non-SMARCA4 non-expressing cells treated with si-SMARCA2.
3 is a diagram confirming the metastatic potential of SMARCA4 in a mouse HCC model:
A: Migration and invasion of si-SMARCA4-treated HCC cells;
B: image of migrating cells;
C: Cell cycle profile of si-SMARCA4-treated HCC cells;
D: Western blot analysis of cell cycle-related proteins in HCC cell lines;
E: si-Cont, si-Smarca4, si-Smarcc1 and pBJ-Smarca2 dosing schedules, ultrasonographic images, and tumor mass and mass number in the Ras-Tg mouse model; and
F: Western blot analysis results of Smarca4, Smarcc1 and Smarca2 in liver tissue of the Ras-Tg mouse model.
4 is a diagram confirming specific target genes regulated by SMARCA4 in HCC:
A: Pie charts, Venn diagrams and schematics of the enhancer gene and SMARCA4-disruption gene;
B: Heatmap of 563 SMARCA4-related genes in multi-stage HCC;
C: functional analysis of 563 genes;
D: Results of SMARCA4 and IRAK1 expression analysis in si-SMARCA4-treated HCC cell lines (qRT-PCR);
E: ChIP-qPCR assay results to evaluate SMARCA4 binding to the IRAK1 enhancer; and
F: Result of dual luciferase analysis in HCC cell line expressing reporter plasmid expressing IRAK1 enhancer region after si-SMARCA4 treatment.
5 is a diagram confirming SMARCA4 and IRAK1 overexpression in the HCC patient cohort.
Figure 6 is a diagram confirming the anti-tumor effect in HCC by target-inhibition of IRAK1:
A: differential gene expression of the IRAK1 gene;
B: Cell growth (cell viability) and cell proliferation rate after SMARCA4 or IRAK1 gene knockdown;
C: cell cycle profile;
D: Western blot analysis of cell cycle-related proteins;
E: Cell images and cell numbers confirmed using Boyden chamber motility analysis; and
F: Cell image and cell number confirmed using transwell invasion assay.
7 is a diagram confirming the regulation of IRAK1 activity of SMARCA4 in HCC:
A: Anti-tumor effect by IRAK1 (pcDNA3.1_IRAK1) after knockdown of SMARCA4 (Western blot analysis, MTT analysis and BrdU analysis);
B: cell cycle profile;
C: The result of confirming the cell cycle regulatory protein by Western blot analysis;
D: Cell images and cell numbers confirmed using Boyden chamber motility analysis; and
E: Cell image and cell number confirmed using transwell invasion assay.
8 is a diagram confirming that SMARCA4 regulates IRAK1 in HCC to activate the oncoproteins Gankyrin and AKR1B10:
A: Western blot analysis of downstream regulatory molecules of IRAK1;
B: Western blot analysis confirmed the expression change of downstream regulatory molecules of IRAK1 by SMARCA4 knockdown (si-SMARCA4) and IRAK1 recovery (pcDNA3.1_IRAK1);
C: dosing schedule of Ras -Tg mouse model;
D: tumor mass number;
E: Western blot analysis of liver tissue from Ras-Tg mice; and
F: Schematic diagram of the mechanism of SMARCA4-IRAK1-Gankyrin, AKR1B10.

Hereinafter, the present invention will be described in detail by way of embodiments of the present invention with reference to the accompanying drawings. However, the following embodiments are presented as examples of the present invention, and when it is determined that detailed descriptions of well-known techniques or configurations known to those skilled in the art may unnecessarily obscure the gist of the present invention, the detailed description may be omitted, and , the present invention is not limited thereby. Various modifications and applications of the present invention are possible within the scope of equivalents interpreted therefrom and the description of the claims to be described later.

In addition, the terms used in this specification are terms used to properly express the preferred embodiment of the present invention, which may vary depending on the intention of a user or operator or customs in the field to which the present invention belongs. Accordingly, definitions of these terms should be made based on the content throughout this specification. Throughout the specification, when a part "includes" a certain component, it means that other components may be further included, rather than excluding other components, unless otherwise stated.

As used herein, the term "subject" or "patient" means any single individual in need of treatment, including humans, apes, monkeys, cattle, dogs, guinea pigs, rabbits, chickens, insects, and the like. Also included in the subject are any subjects who participated in a clinical study trial without any clinical manifestations of any disease, or subjects who participated in epidemiological studies or subjects used as controls.

As used herein, the term “sample (sample)” refers to a biological sample obtained from a subject or patient. Sources of biological samples may include fresh, frozen and/or preserved organ or tissue samples or solid tissue from biopsies or aspirates; blood or any blood component; The cells may be at any point in the pregnancy or development of the subject.

All technical terms used in the present invention, unless otherwise defined, have the meaning as commonly understood by one of ordinary skill in the art of the present invention. In addition, although preferred methods and samples are described herein, similar or equivalent ones are also included in the scope of the present invention. The contents of all publications herein incorporated by reference are incorporated herein by reference.

In one aspect, the present invention is selected from the group consisting of SMARCA4 (SWI / SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 4), SMARCC1, SMARCA2 and IRAK1 (Interleukin-1 receptor-associated kinase 1) It relates to a biomarker for diagnosing liver cancer comprising one or more genes or proteins expressed from the genes.

In one embodiment, the biomarker may include genes of SMARCA4 and IRAK1 or proteins expressed from the genes together.

In one embodiment, the liver cancer may be hepatocellular carcinoma (HCC), more preferably hepatocellular carcinoma overexpressing IRAK1.

In one embodiment, it may further include a gene of Gankyrin or AKR1B10 (ldo-keto reductase family 1 member B10) or a protein expressed from the gene.

In one embodiment, SMARCA4, SMARCC1, IRAK1, Gankyrin or AKR1B10 may increase liver cancer-specific expression, and SMARCA2 may decrease liver cancer-specific expression.

In one aspect, the present invention relates to a composition for diagnosing liver cancer, comprising an agent for measuring the expression level of one or more biomarker genes selected from the group consisting of SMARCA4, SMARCC1, SMARCA2 and IRAK1 at the mRNA or protein level.

In one embodiment, the composition may further comprise an agent for measuring the expression level of Gankyrin or AKR1B10 at the mRNA or protein level.

In one embodiment, the liver cancer may be hepatocellular carcinoma, more preferably hepatocellular carcinoma overexpressing IRAK1.

In one embodiment, the composition of the present invention may include an agent for measuring the expression levels of SMARCA4 and IRAK1 genes at the mRNA or protein level.

In one embodiment, the agent for measuring the expression level of a biomarker gene at the mRNA level is a nucleic acid sequence of the marker, a nucleic acid sequence complementary to the nucleic acid sequence, a fragment of the nucleic acid sequence and the complementary sequence to specifically recognize It may include a primer pair, a probe, or a primer pair and a probe, and the measurement is a polymerase chain reaction, real-time RT-PCR, reverse transcription polymerase chain reaction, competitive polymerase chain reaction ( Competitive RT-PCR), nuclease protection assay (RNase, S1 nuclease assay), in situ hybridization, nucleic acid microarray, Northern blot, or a method selected from the group consisting of a DNA chip.

In one embodiment, the agent for measuring the expression level of a biomarker gene at the protein level is an antibody, antibody fragment, aptamer, avidity multimer that specifically recognizes the full protein length of the marker or a fragment thereof ) or peptidomimetics, and the measurement may be Western blot, ELISA (enzyme linked immunosorbent assay), radioimmunoassay (RIA), radioimmunodiffusion, immunoelectrophoresis, tissue immunostaining , immunoprecipitation assay, complement fixation assay, FACS, mass spectrometry, or a method selected from the group consisting of protein microarray.

As used herein, the term “detection” or “measurement” refers to quantifying the concentration of a detected or measured target.

As used herein, the term "primer" is a nucleic acid sequence having a short free 3 hydroxyl group, which can form a complementary template and base pair, and serves as a starting point for template strand copying. It refers to a short nucleic acid sequence that functions. The primers can initiate DNA synthesis in the presence of reagents for polymerization (ie, DNA polymerate or reverse transcriptase) and the four different nucleoside triphosphates in an appropriate buffer and temperature.

As used herein, the term "probe" refers to a nucleic acid fragment such as RNA or DNA corresponding to several bases to several hundred bases as short as possible to achieve specific binding to mRNA, and is labeled to determine the presence or absence of a specific mRNA. can The probe may be manufactured in the form of an oligonucleotide probe, a single stranded DNA probe, a double stranded DNA probe, an RNA probe, or the like. In the present invention, hybridization is performed using a probe complementary to the AFP, HMMR, NXPH4, PITX1, THBS4 and/or UBE2T genes, and the level of gene expression can be diagnosed based on whether hybridization occurs. The selection of suitable probes and hybridization conditions may be modified based on those known in the art, and therefore, the present invention is not particularly limited thereto.

The primers or probes of the present invention can be chemically synthesized using the phosphoramidite solid support method, or other well-known methods. Such nucleic acid sequences may also be modified using a number of means known in the art. Non-limiting examples of such modifications include methylation, encapsulation, substitution with one or more homologues of natural nucleotides, and modifications between nucleotides, such as uncharged linkages such as methyl phosphonates, phossotriesters, phosphoro amidates, carbamates, etc.) or charged linkages (eg phosphorothioates, phosphorodithioates, etc.).

In the present invention, suitable conditions for hybridizing a probe with a cDNA molecule can be determined in a series of processes by an optimization procedure. These procedures are carried out as a series of procedures by those skilled in the art to establish protocols for use in the laboratory. For example, conditions such as temperature, concentration of components, hybridization and washing times, buffer components and their pH and ionic strength depend on various factors such as probe length and GC amount and target nucleotide sequence. Detailed conditions for hybridization are described in Joseph Sambrook, et al., Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (2001); and M.L.M. Anderson, NucleicAcidHybridization, Springer-Verlag New York Inc. N.Y. (1999). For example, high stringency among the stringent conditions is 0.5 M NaHPO4, 7% sodium dodecyl sulfate (SDS), 1 mM EDTA at 65° C., and 0.1×SSC (standard saline citrate)/0.1% SDS at 68° C. It means washing under conditions. Alternatively, high stringency conditions mean washing at 48° C. in 6×SSC/0.05% sodium pyrophosphate. Low stringency conditions mean washing at 42° C. in, for example, 0.2×SSC/0.1% SDS.

As used herein, the term “antibody” refers to a specific protein molecule directed to an antigenic site as a term known in the art. For the purpose of the present invention, an antibody refers to an antibody that specifically binds to a protein expressed in AFP, HMMR, NXPH4, PITX1, THBS4 and/or UBE2T genes, which are markers of the present invention, and the method for preparing the antibody is widely It can be prepared using a known method. This also includes partial peptides that can be made from the protein. The form of the antibody of the present invention is not particularly limited, and a part thereof is also included in the antibody of the present invention as long as it has a polyclonal antibody, a monoclonal antibody, or antigen-binding property, and all immunoglobulin antibodies are included. Furthermore, the antibody of the present invention includes a special antibody such as a humanized antibody.

In one aspect, the present invention relates to a liver cancer diagnostic kit comprising the composition of the present invention.

In one embodiment, the kit may further include tools and/or reagents for collecting a biological sample from a subject or patient, as well as tools and/or reagents for preparing genomic DNA, cDNA, RNA or protein from the sample. have. For example, it may include PCR primers for amplifying a relevant region of genomic DNA. The kit may include probes of genetic factors useful for pharmacogenomic profiling. In addition, in the use of such a kit, the labeled oligonucleotide can be easily identified during analysis.

In the present invention, the term “diagnosis” as used herein refers to determining the susceptibility of an object to a specific disease or disorder, determining whether an object currently has a specific disease or disorder, a specific disease or disorder Determining the prognosis of a subject with a disease (eg, identification of a pre-metastatic or metastatic cancer state, staging the cancer, or determining the responsiveness of the cancer to treatment), or therametrics (eg, monitoring the condition of the subject to provide information on treatment efficacy).

In one embodiment, the kit may further contain a labeling material such as a DNA polymerase and dNTPs (dGTP, dCTP, dATP and dTTP), a fluorescent material, and the like.

In one aspect, the present invention relates to a pharmaceutical composition for preventing or treating liver cancer comprising a SMARCA4 inhibitor as an active ingredient.

In one embodiment, the SMARCA4 inhibitor may be siRNA, or siRNA of SEQ ID NO: 1.

In one embodiment, SMARCA2 or an expression promoter thereof, a Gankyrin inhibitor, an AKR1B10 inhibitor, a SMARCC1 inhibitor, or an IRAK1 inhibitor may be further included.

In one embodiment, the IRAK1 inhibitor may be siRNA, or siRNA of SEQ ID NO: 2.

In one embodiment, the liver cancer may be hepatocellular carcinoma, more preferably hepatocellular carcinoma overexpressing IRAK1.

The pharmaceutical composition according to the present invention may include any substance capable of inhibiting the expression of SMARCA4, SMARCC1, or IRAK1 or promoting the expression of SMARCA2.

In the present invention, the pharmaceutical composition of the present invention may further include a pharmaceutically acceptable carrier. As used herein, "pharmaceutically acceptable" refers to a composition that is physiologically acceptable and does not normally cause allergic reactions or similar reactions such as gastrointestinal disorders and dizziness when administered to humans. Pharmaceutically acceptable carriers include, for example, carriers for oral administration such as lactose, starch, cellulose derivatives, magnesium stearate, stearic acid, and the like, and carriers for parenteral administration such as water, suitable oils, saline, aqueous glucose and glycols, and the like. and the like, and may further include a stabilizer and a preservative. Suitable stabilizers include antioxidants such as sodium hydrogen sulfite, sodium sulfite or ascorbic acid. Suitable preservatives are benzalkonium chloride, methyl- or propyl-paraben and chlorobutanol. As other pharmaceutically acceptable carriers, reference may be made to those described in the following literature (Remington's Pharmaceutical Sciences, 19th ed., Mack Publishing Company, Easton, PA, 1995).

The pharmaceutical composition according to the present invention may be formulated in a suitable form according to a method known in the art together with a pharmaceutically acceptable carrier as described above. That is, the pharmaceutical composition of the present invention can be prepared in various parenteral or oral dosage forms according to known methods, and as a representative dosage form for parenteral administration, an isotonic aqueous solution or suspension is preferred as an injectable dosage form. Formulations for injection may be prepared according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. For example, each component may be formulated for injection by dissolving it in saline or buffer. In addition, formulations for oral administration include, but are not limited to, powders, granules, tablets, pills and capsules. The pharmaceutical composition formulated in the above manner may be administered in an effective amount through various routes including oral, transdermal, subcutaneous, intravenous or intramuscular, and the “administration” refers to introducing a predetermined substance into a patient by any suitable method. and the route of administration of the substance can be administered through any general route as long as it can reach the target tissue. As used herein, the term “effective amount” refers to an amount exhibiting a preventive or therapeutic effect when administered to a patient. The dosage of the pharmaceutical composition according to the present invention may vary depending on various factors such as the type and severity of the patient's disease, age, sex, weight, sensitivity to drugs, the type of current treatment, administration method, target cell, etc. can be easily determined by experts in In addition, the pharmaceutical composition of the present invention may be administered in combination with a conventional therapeutic agent, may be administered sequentially or simultaneously with the conventional therapeutic agent, and may be administered single or multiple. Preferably, in consideration of all of the above factors, an amount capable of obtaining the maximum effect with a minimum amount without side effects can be administered, more preferably from 1 to 10000 μg/weight kg/day, even more preferably from 10 to 1000 It can be administered repeatedly several times a day at an effective dose of mg/kg body weight/day.

In one embodiment, the SMARCA2 expression promoter, preferably a chemical substance, a nucleotide, a vector containing the gene, a protein or fragment thereof into which the gene is translated, or a natural product extract may be included as an active ingredient.

As used herein, the term "promotion of expression" means to cause enhancement of expression (to mRNA) or translation (to protein) of a target gene.

In one embodiment, the expression inhibitor of SMARCC1, IRAK1, Gankyrin or AKR1B10, preferably a chemical substance, nucleotide, antisense, siRNA oligonucleotide or a natural product extract as an active ingredient, more preferably, the nucleotide of the gene An antisense or small interference RNA (siRNA) oligonucleotide having a sequence complementary to the sequence may be included as an active ingredient.

As used herein, the term "repression of expression" means to cause a decrease in expression (to mRNA) or translation (to protein) of a target gene, preferably by which the target gene expression becomes undetectable or insignificant. means to exist as

As used herein, the term "small interfering RNA (siRNA)" refers to a nucleic acid molecule capable of mediating RNA interference or gene silencing (International Patent Publication Nos. 00/44895, 01/36646, 99/32619, 01 see /29058, 99/07409 and 00/44914). Since siRNA can inhibit the expression of a target gene, it is provided as an efficient gene knock-down method or as a gene therapy method. In the siRNA molecule of the present invention, the sense strand (sequence corresponding to the mRNA sequence of SMARCC1, IRAK1, Gankyrin or AKR1B10) and the antisense strand (sequence complementary to the mRNA sequence of SMARCC1, IRAK1, Gankyrin or AKR1B10) are located opposite to each other. It may have a double-stranded structure, and further, the siRNA molecule of the present invention may have a single-stranded structure having self-complementary sense and antisense strands. Furthermore, siRNA is not limited to the complete pairing of double-stranded RNA parts that are paired with each other, but pairs are formed by mismatch (corresponding bases are not complementary), bulges (there is no base corresponding to one chain), etc. There may be parts that are not achieved. In addition, as long as the siRNA end structure can suppress the expression of SMARCC1, IRAK1, Gankyrin or AKR1B10 genes by the RNAi effect, both blunt ends or cohesive ends are possible, and the cohesive end structure is 3'-end protrusion Both structures and 5'-end overhang structures are possible. In addition, the siRNA molecule of the present invention may have a form in which a short nucleotide sequence is inserted between the self-complementary sense and antisense strands, and in this case, the siRNA molecule formed by expression of the nucleotide sequence is used for intramolecular hybridization Thus, a hairpin structure is formed, and as a whole, a stem-and-loop structure is formed (shRNA). This stem-and-loop structure can be processed in vitro or in vivo to generate active siRNA molecules capable of mediating RNAi. Methods for preparing siRNA include direct synthesis of siRNA in a test tube and introduction into cells through transformation, and transfection of an siRNA expression vector or PCR-derived siRNA expression cassette prepared so that siRNA is expressed in the cell. There is a method of converting or infecting.

In one embodiment, the composition of the present invention comprising a gene-specific siRNA may include an agent that promotes the influx of the siRNA into a cell. In general, an agent that promotes nucleic acid influx can be used for the agent that promotes the influx of siRNA into the cell. For example, a parent of one of a number of sterols including cholesterol, cholate and deoxycholic acid using liposomes. It may be formulated with an oily carrier. Also poly-L-lysine (poly-L-lysine), spermine (spermine), polysilazane (polysilazane), polyethylenimine (PEI: polyethylenimine), polydihydroimidazolenium (polydihydroimidazolenium), polyallylamine (polyallylamine) ), a cationic polymer such as chitosan may be used, and succinylated PLL (succinylated PLL), succinylated PEI (succinylated PEI), polyglutamic acid, polyaspartic acid (polyaspartic acid), polyacrylic acid (polyacrylic acid), polymethacylic acid (polymethacylic acid), dextran sulfate (dextran sulfate), heparin (heparin), anionic polymers such as hyaluronic acid (hyaluronic acid) Available.

In one embodiment, when an antibody specific for the protein is used as a substance for reducing the expression and activity of the SMARCC1, IRAK1, Gankyrin or AKR1B10 protein, the antibody is directly or indirectly coupled to an existing therapeutic agent or through a linker. (eg, covalently).

As used herein, the term "antisense oligonucleotide" refers to DNA or RNA or a derivative thereof containing a nucleic acid sequence complementary to a specific mRNA sequence, and is bound to a complementary sequence in mRNA to form SMARCC1, IRAK1, Gankyrin or acts to inhibit the translation of AKR1B10 into protein.Antisense sequence of the present invention is complementary to SMARCC1, IRAK1, Gankyrin or AKR1B10 mRNA and refers to a DNA or RNA sequence capable of binding to SMARCC1, IRAK1, Gankyrin or AKR1B10 mRNA and inhibits the essential activity of SMARCC1, IRAK1, Gankyrin or AKR1B10 mRNA translation, translocation into the cytoplasm, maturation or all other overall biological functions.In addition, the antisense nucleic acid enhances efficacy It may be modified at one or more bases, sugars, or backbone positions to allow it to be modified (De Mesmaeker et al., Curr Opin Struct Biol., 5, 3, 343-55, 1995). acid, phosphotriester, methyl phosphonate, short chain alkyl, cycloalkyl, short chain heteroatomic, heterocyclic intersugar linkage, etc. Antisense nucleic acids may also contain one or more substituted sugar moieties. Antisense nucleic acid may comprise modified bases.Modified bases include hypoxanthine, 6-methyladenine, 5-methylpyrimidine (especially 5-methylcytosine), 5-hydroxymethylcytosine. (HMC), glycosyl HMC, gentobiosyl HMC, 2-aminoadenine, 2-thiouracil, 2-thiothymine, 5-bromouracil, 5-hydroxymethyluracil, 8-azaguanine, 7-de azaguanine, N6 (6-aminohexyl) adenine, 2,6-diaminopurine, etc. In addition, the antisense nucleic acid of the present invention is effective in reducing the activity and It may be chemically bound to one or more moieties or conjugates that enhance fabric adsorption properties. Cholesterol moiety, cholesteryl moiety, cholic acid, thioether, thiocholesterol, fatty chain, phospholipid, polyamine, polyethylene glycol chain, adamantane acetic acid, palmityl moiety, octadecylamine, hexylaminocarbonyl-oxychol fat soluble moieties such as esterol moieties, and the like. Oligonucleotides comprising a fat-soluble moiety and methods of preparation are well known in the art (US Pat. Nos. 5,138,045, 5,218,105 and 5,459,255). The modified nucleic acid may increase the stability to nucleases and increase the binding affinity between the antisense nucleic acid and the target mRNA. Antisense oligonucleotides can be synthesized in vitro by a conventional method and administered in vivo, or antisense oligonucleotides can be synthesized in vivo. An example of synthesizing antisense oligonucleotides in vitro is using RNA polymerase I. One example of allowing antisense RNA to be synthesized in vivo is to use a vector with the origin of the recognition site (MCS) in the opposite direction to allow the antisense RNA to be transcribed. Such antisense RNA preferably has a translation stop codon in the sequence so that it is not translated into the peptide sequence.

In one aspect, the present invention comprises the steps of (a) measuring the gene expression level of SMARCA4 or IRAK1 in a biological sample isolated from a test subject; (b) comparing the corresponding gene of the normal control sample with the corresponding result; and (c) when the gene expression level in step (a) is higher than the expression level of the corresponding gene in step (b), determining that the test subject is liver cancer, providing information necessary for diagnosing liver cancer It's about how to do it.

In one embodiment, the method comprises the steps of (a) measuring the gene or protein expression level of SMARCC1, SMARCA2, Gankyrin, or AKR1B10 in a biological sample isolated from a test subject; (b) comparing the corresponding gene or protein of the normal control sample with the corresponding result; And (c) when the expression level in step (a) of SMARCC1, Gankyrin or AKR1B10 is high compared to the expression level of the corresponding gene or protein in step (b), or the expression level in step (a) of SMARCA2 (b) When it is low compared to the expression level of the gene or protein in the step, the step of determining that the test subject is liver cancer may be further included.

In one embodiment, the biological sample may include a sample such as tissue, cells, whole blood, serum, plasma, saliva, sputum, cerebrospinal fluid or urine, and the like.

In one aspect, the present invention comprises the steps of (a) measuring the gene expression level of SMARCA4 or IRAK1 in a biological sample isolated from a test subject; (b) comparing the corresponding gene of the normal control sample with the corresponding result; and (c) when the gene expression level in step (a) is higher than the expression level of the corresponding gene in step (b), determining that the prognosis of the test subject is bad, for predicting the prognosis of liver cancer How to provide information.

In one aspect, the present invention relates to a method for predicting liver cancer treatment responsiveness of a SMARCA4 inhibitor, comprising measuring the expression level of IRAK1.

In one embodiment, the method may further comprise measuring the expression level of Gankyrin or AKR1B10.

The present invention will be described in more detail through the following examples. However, the following examples are only intended to embody the contents of the present invention, and the present invention is not limited thereto.

Example 1. Evaluation of subunit gene expression level of SWI/SNF complex in HCC

To confirm the expression level of subunit genes in HCC, TCGA_LIHC (The Cancer Genome Atlas liver hepatocellular carcinoma project), ICGC_LIRI (International Cancer Genome Consortium liver Cancer-RIKEN, JP) and NCBI's Gene Expression Omnibus (GEO) database (Accession) Numbers: GSE6764, GSE22058, GSE25097, GSE54238, GSE62043, GSE77314, GSE89377 and GSE114564). The level 3 mRNA expression data of TCGA-LIHC HTSeq-FPKM were log2 transformed and [log2(fpkm+1)] used to evaluate gene expression levels. As a result, it was found that human HCC (cBioPortal; n=1,019) had a very low mutation rate of the SWI/SNF subunit gene ( FIG. 1A ). The low incidence and heterogeneity of such genetic alterations suggests that the predominantly aberrant expression of specific subunit genes may cause systemic disturbances of the SWI/SNF complex. Accordingly, as a result of examining the expression of SWI/SNF subunit genes using multi-step HCC data (Catholic_mLIHC; GSE114564) and TCGA_LIHC, ICGC_LIRI, GSE77314, GSE22058, GSE25097, GSE62043 and GSE6764 data sets ( FIG. 1B ), 10 SWI/ Among the SNF subunit genes, SMARCA4 and SMARCC1 were significantly overexpressed in HCC patients, and it was confirmed that SMARCA2 was down-regulated at the same time (≥ ± 1.5 fold, P < 0.05). In addition, as a result of analyzing 34 HCC tissues and corresponding HCC tissue pairs from noncancerous liver biopsies obtained from the National Biobank of Korea, abnormal regulation of SMARCA4, SMARCA2 and SMARCC1 was consistent in TCGA_LIHC, ICGC_LIRI and GSE77314. It was also identified in tissue pairs of HCC (Fig. 1C). In addition, further validation with qRT-PCR in an additional set of 34 randomly selected HCC tissues, SMARCA4 (67.6%) and SMARCC1 (52.9%) overexpression and SMARCA2 (61.8%) inhibition were consistently observed in this set ( 1D). In addition, as a result of analyzing 10 tissue pairs between HCC and noncancerous liver selected by Western blot, significant overexpression of SMARCA4 and SMARCC1 and inhibition of SMARCA2 were observed in HCC at the same time ( FIG. 1E ).

Example 2. Anti-tumor effect by regulation of expression of SMARCA4, SMARCA2 and SMARCC1

2-1. in vitro analysis

In order to confirm the role of SMARCA4, SMARCA2, and SMARCC1 whose HCC-specific expression is changed in the above example in hepatocarcinogenesis, each gene was knocked down by RNA interference. To this end, each gene was knocked down by RNA interference in Hep3B, Huh7 and SNU-449 liver cell lines with relatively high SMARCA4 and SMARCC1 expression and relatively low SMARCA2 expression than in normal hepatocytes, and cell viability and cell proliferation rates were analyzed by MTT analysis, BrdU Assays and colonogenic assays were confirmed. In addition, the migration and invasion of knock-down HCC cells of each gene were confirmed by Boyden chamber motility assay and wound healing assay. In addition, the effect of knockdown of each gene on cell cycle regulation in HCC cells was confirmed by flow cytometric analysis, and cell cycle regulatory proteins were confirmed by Western blot analysis.

As a result, it was shown that the knockdown of SMARCA4 and SMARCC1 significantly inhibited the cell growth and proliferation rate of HCC cells ( FIGS. 2A to 2D ). In addition, in both SMARCA4-null HCC cell lines PLC/PRF/5 and SK-Hep-1, SMARCA2 knockdown resulted in overall HCC cell lethality, whereas SMARCA4 high-expressing Hep3B cells and Huh7 cells showed a change in growth rate. was found to be absent (Fig. 2E). These results indicate a pivotal role of the SMARCA4-SWI/SNF complex in the development of hepatocellular carcinoma. In addition, the results of the Boyden chamber motility assay showed that SMARCA4 knockdown significantly inhibited the serum-stimulated migration and invasion response of HCC cells (FIG. 3A), and similarly, wound healing of HCC cells in the wound healing assay results. The effect was shown to be reduced by SMARCA4 knockdown ( FIG. 3B ). In addition, as a result of confirming the effect of SMARCA4 knockdown on the cell cycle regulation of HCC cells by flow cytometry, it was found that SMARCA4 knockdown increases the number of cells in the G1 phase in HCC cells ( FIG. 3C ), and cell cycle by Western blot As a result of confirming the expression of regulatory proteins, SMARCA4 knockdown selectively induces p27 Kip1 expression in HCC cells, and at the same time inhibits cyclin D1, cyclin E, cyclin-dependent kinase 2 (CDK2), CDK4 and CDK6, thereby hyperphosphorylating pRb was shown to induce (hypophosphorylation) (p-pRb) ( FIG. 3D ). Through this, it was found that SMARCA4 hyperactivity in HCC regulates the electrons of cell cycle proteins and promotes the conversion from G1 to S phase.

2-2. in vivo analysis

To determine whether the regulation of SMARCA4, SMARCA2, and SMARCC1 induces tumor suppressive effects in vivo, SMARCA4 from 14 weeks of age in 8 Ras-Tg (H-ras12V homozygous transgenic) male mice, in which HCC develops at about 15 to 18 weeks of age. - Liver-specific delivery reagent Invivofectamine mixed with target mouse siRNA (si-Smarca4) or SMARCC1 siRNA (si-Smarcc1), or Turbofect mixed with mouse SMARCA2 expression plasmid (pBJ-Smarca2) was intravenously injected (Fig. 3E). Liver HCC of mice was confirmed by ultrasonography (Philips, Amsterdam, Nederland) from 14 weeks of age to 24 weeks of age to obtain a liver, and the obtained liver weight and tumor mass number were confirmed and disrupted by Western blot analysis SMARCA4, SMARCA2 and SMARCC1 was confirmed.

As a result, in control mice (si-Cont), HCC was detected from the age of 18 weeks, and 3 out of 4 mice developed many large HCCs. On the other hand, no HCC was detected in the mice of the si-Smarca4 treatment group until 24 weeks of age. However, HCC was detected at 20 weeks of age in si-Smarcc1 treated mice and pBJ-Smarca2 treated mice, and relatively small HCC was developed in 3 out of 4 mice. Total weight change and tumor mass number of the liver indicated that target-inhibition of SMARCA4 was remarkably effective in reducing tumor burden. In addition, as a result of Western blot analysis, suppression of Smarca4 and Smarcc1 and increased expression of Smarca2 were confirmed in the mouse liver tissues of each group ( FIG. 3F ).

Example 3. Identification of SMARCA4-regulated genes in HCC

3-1. SMARCA4-regulated gene selection

To identify SMARCA4-SWI/SNF regulatory genes, by SMARCA4 in HCC cells using histone ChIP-seq of HepG2 cells available from ENCODE (Encyclopedia of DNA Elements) (https://www.encodeproject.org/) Locations rich in specifically recognized markers, H3K27ac and H3K27me3, were comprehensively mapped. Trimmed reads were aligned to the human genome using the Bowtie 2 ( http://bowtie-bio.sourceforge.net/bowtie2/index.shtml) mapper here, and duplicate reads were used to eliminate PCR bias. The values were removed using the Sambamba tool ( https://lomereiter.github.io/sambamba/ ). H3K27ac or H3K27me3 rich sites were identified using HOMER (findPeaks) (FDR-adjusted p-value cutoff=0.001). Through the above analysis, 9,198 genes out of 21,684 genes closest to the peak were identified as active enhancers (FIG. 4A). Then, to identify the regulated gene elements in HCC cells, SMARCA4 knockdown was performed in Hep3B to find 1,865 gene elements regulated by SMARCA4 (> ± 1.5 fold, P < 0.05), of which 1,054 genes were shown to be downregulated in SMARCA4 knockdown HCC cells. As a result of analyzing 1,054 SMARCA4-regulated genes together with 9,198 active enhancers by Venn diagram analysis, 563 genes were analyzed to be regulated by SMARCA4-SWI/SNF in HCC (FIG. 4A). 563 SMARCA4-associated genes showed progressive increases or decreases in transcription levels across different stages of HCC, and consistently highest or lowest levels in advanced HCC ( FIG. 4B ). Using Gene Ontology terms, genes related to carcinogenesis functions including angiogenesis, Wnt signaling, integrin signaling, PDGF signaling and G-protein signaling were searched for in the 563 gene sets (Fig. 4C). ). To select for genes regulated by SMARCA4 in HCC, 563 genes were reduced to 84 genes overexpressed in both TCGA_LIHC and ICGC_LIRI data sets (≥ 1.5 fold, P < 0.05), and then significantly correlated with SMARCA4 expression Among the 37 genes of interest, the top 5 genes that were upregulated were tested and validated ( FIG. 4A ). To confirm whether SMARCA4 directly regulates these genes, HCC cells were knocked down with SMARCA4 siRNA (si-SMARCA4) and gene expression changes were confirmed by qRT-PCR. As a result, it was confirmed that CKAP4 and IRAK1 were consistently inhibited by SMARCA4 knockdown in all HCC cells ( FIG. 4D ).

3-2. Confirm the interaction of SMARCA4 and IRAK1

To confirm that SMARCA4 specifically binds to the enhancer regions of the two genes CKAP4 and IRAK1 selected in the above example, ChIP analysis was performed using anti-SMARCA4 and anti-H3K27ac antibodies, respectively, and the active enhancer regions of CKAP4 and IRAK1 Enrichment rates in (H3K27ac) were confirmed. Chromatin immunoprecipitation (ChIP) analysis was performed according to the manufacturer's (Pierce Agarose ChIP kit; Thermo Fisher Scientific, Waltham, MA) instructions, and DNA was RT- with primers for the enhancer region of each candidate gene regulated by SMARCA4. It was amplified by qPCR. Cross-linked DNA was precipitated using SMARCA4 antibody (anti-SMARCA4) and acetylated histone H3 at lysine 27 (H3K27ac) antibody (Abcam, Cambridge, UK) (anti-H3K27ac). The experiment was repeated three times and the average was obtained by normalizing to IgG. In addition, to confirm that endogenous SMARCA4 directly binds to the IRAK1 enhancer region, after cloning the IRAK1 enhancer into the pGL4.23 reporter vector, a dual luciferase reporter assay of the pGL4.23_IRAK1 enhancer in the presence or absence of SMARCA4 in HCC cells was performed. carried out.

As a result, the enrichment rate of the IRAK1 activity enhancer region by SMARCA4 knockdown in all HCC cells was significantly reduced compared to the negative control (randomly selected non-SMARCA4 regulatory genes GNLS, EIF4G2 and CCT2), but the CKAP4 activity enhancer in the same cells Regions were not consistently affected by SMARCA4 knockdown ( FIG. 4E ). In addition, as a result of the luciferase reporter analysis, SMARCA4 knockdown in all HCC cells significantly inhibited the relative luciferase activity, confirming the interaction between SMARCA4 and the IRAK1 enhancer ( FIG. 4F ).

Example 4. Confirmation of SMARCA4 and IRAK1 overexpression in HCC patient cohort

As a result of confirming the overexpression of SMARCA4 and IRAK1 in the TCGA_LIHC and ICGC_LIRI data sets, in TCGA_LIHC, among a total of 371 HCC patients, 318 patients with increased SMARCA4 expression at least 1.5 times the average SMARCA4 or IRAK1 gene expression value of 50 normal patients. (86%), and 335 patients (90%) had increased IRAK1 expression, of which 304 out of 318 patients (96%) had both SMARCA4 and IRAK1 overexpression. (Fig. 5). In addition, in ICGC_LIRI, among a total of 238 HCC patients, 172 patients (72%) with increased SMARCA4 expression by 1.5 times or more of the average SMARCA4 or IRAK1 gene expression value of 202 normal patients, and those with increased IRAK1 expression The number of patients was 198 (83%), of which 152 (88%) of 172 patients were both SMARCA4 and IRAK1 overexpressed at the same time (FIG. 5). Through this, it can be inferred that the expression of IRAK1 is increased by SMARCA4.

Example 5. Identification of the tumorigenic role of IRAK1 in HCC

Since SMARCA4 activated IRAK1 in HCC cells through the above examples, in order to confirm the role of IRAK1 in hepatocellular carcinogenesis, the expression change of IRAK1 in TCGA_LIHC and ICGC_LIRI was confirmed. As a result, SMARCA4 of IRAK1 A strong positive correlation was observed with (Fig. 6A). In addition, the Kaplan-Meier survival curve of HCC patients showed that the 5-year overall survival rate of patients with high IRAK1 expression was significantly lower than that of patients with low IRAK1 expression ( FIG. 6A ). Therefore, in order to confirm the tumorigenic function of IRAK1 in the development of hepatocellular carcinoma, cell growth using MTT assay and cell proliferation using BrdU assay were confirmed in HCC cells. As a result, individual knockdown of SMARCA4 and IRAK1 significantly inhibited tumor cell growth and proliferation in HCC cells ( FIG. 6B ). In addition, as a result of confirming the PI-stained cells by flow cytometry to confirm cell cycle regulation, it was shown that each knockdown of SMARCA4 and IRAK1 induced cell cycle arrest of HCC cells. In particular, cell cycle components regulated by SMARCA4 were regulated together with IRAK1, confirming that IRAK1 is a downstream molecule regulated by SMARCA4 in HCC ( FIGS. 6C and D ). In addition, the knockdown of SMARCA4 and IRAK1 reduced the migratory and invasion responses of chemoattractant-stimulated HCC cells ( FIGS. 6E and F ).

Example 6. Regulation of IRAK1 activity by SMARCA4

To confirm whether SMARCA4 directly regulates IRAK1 activity, HCC cells were knocked down with SMARCA4 siRNA, and then a recovery experiment was performed with an IRAK1 expression plasmid (pcDNA3.1_IRAK1). As a result, SMARCA4 knockdown significantly inhibited both tumor cell growth and proliferation of HCC cells, but this anti-tumor effect was abolished by co-transformation of the IRAK1 plasmid ( FIG. 7A ). In addition, as a result of flow cytometry analysis of PI-stained cells, SMARCA4 knockdown in HCC cells significantly induced G1 phase arrest, showing anti-growth effects, whereas co-transformation of the same cells with IRAK1 plasmid resulted in HCC cell cycle It was not stopped and tumor growth resumed again ( FIG. 7B ). These results were further verified by confirming cell cycle-related proteins by Western blot analysis. As a result, p27 Kip1 selectively induced by SMARCA4 knockdown in HCC was inhibited by IRAK1 plasmid expression, and at the same time by SMARCA4 knockdown. Phosphorylation of the suppressed cyclin D1, cyclin E, CDK2, CDK4 and CDK6 and pRb was restored ( FIG. 7C ). In addition, chemotactic-stimulated migration and invasion responses reduced by SMARCA4 knockdown in HCC cells were also restored due to the expression of IRAK1 plasmid ( FIGS. 7D and E).

Example 7. Oncoprotein regulation of IRAK1

To confirm whether tumor proteins are regulated by SMARCA4-IRAK1, SMARCA4 and IRAK1 were knocked down in HCC cells, and IRAK1 expression was restored with IRAK1 plasmid to confirm the degree of regulation of tumor proteins. As a result, as a result of knockdown of both SMARCA4 and IRAK1 in HCC, the JNK-dependent Gankyrin and AKR1B10 were significantly inhibited ( FIG. 8A ). In addition, the inhibitory effect was restored by co-expression of IRAK1 using the IRAK1 plasmid, confirming the regulatory axis of SMARCA4-IRAK1-Gankyrin/AKR1B10 in the development of hepatocellular carcinoma ( FIG. 8B ). In addition, to determine whether the in vivo SMARCA4-IRAK1 regulatory axis can be used as a cancer prevention target, Invivofectamine 3.0 (Invitrogen) containing 25 mg/kg of si-Smarca4 or si-Irak1 was administered intravenously to 14-week-old Ras-Tg mice. Injections and ultrasonography at 15, 17, 19, 21 and 23 weeks of age ( FIG. 8C ). As a result, in the control group (si-Cont), HCC was detected at 19 weeks of age, and a large number of large tumors occurred in 4 out of 4 mice. On the other hand, in the si-Irak1-treated group, HCC was observed at 21 weeks of age, and only 1 out of 4 animals developed relatively small HCC, and in the si-Smarca4-treated group, no HCC was detected for 24 weeks. When both SMARCA4 and IRAK1 were targeted, tumor burden was reduced, and total liver weight and tumor mass number were also significantly suppressed ( FIG. 8D ). In addition, in Western blot analysis, the expression of gankyrin and Akr1b10 in the mouse liver was suppressed in the si-Smarca4 and si-Irak1-treated groups, demonstrating an in vivo regulatory mechanism through SMARCA4-IRAK1-Gankyrin and AKR1B10 in the development of hepatocellular carcinoma ( 8E and F).

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Claims (23)

At least one gene or the gene selected from the group consisting of SMARCA4 (SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 4), SMARCC1, SMARCA2, and IRAK1 (Interleukin-1 receptor-associated kinase 1) A biomarker for diagnosing liver cancer comprising a protein expressed from The biomarker for diagnosing liver cancer according to claim 1, comprising the genes of SMARCA4 and IRAK1 or proteins expressed from the genes. The biomarker for diagnosing liver cancer according to claim 1, wherein the liver cancer is hepatocellular carcinoma (HCC). The biomarker for diagnosing liver cancer according to claim 3, which is hepatocellular carcinoma in which IRAK1 is overexpressed. The biomarker for diagnosing liver cancer according to claim 1, further comprising a gene of Gankyrin or AKR1B10 or a protein expressed from the gene. A composition for diagnosing liver cancer, comprising an agent for measuring the expression level of one or more biomarker genes selected from the group consisting of SMARCA4, SMARCC1, SMARCA2 and IRAK1 at the mRNA or protein level. The composition for diagnosing liver cancer according to claim 6, further comprising an agent for measuring the expression level of Gankyrin or AKR1B10 at the mRNA or protein level. The composition for diagnosing liver cancer according to claim 6, wherein the liver cancer is hepatocellular carcinoma. The composition for diagnosing liver cancer according to claim 6, comprising an agent for measuring the expression levels of SMARCA4 and IRAK1 genes at the mRNA or protein level. The method according to claim 6, wherein the agent for measuring the expression level of the biomarker gene at the mRNA level specifically recognizes the nucleic acid sequence of the marker, a nucleic acid sequence complementary to the nucleic acid sequence, and a fragment of the nucleic acid sequence and the complementary sequence which is a primer pair, a probe, or a primer pair and a probe, a composition for diagnosing liver cancer. 11. The method of claim 10, wherein the measurement is polymerase chain reaction, real-time RT-PCR (Real-time RT-PCR), reverse transcription polymerase chain reaction, competitive polymerase chain reaction (Competitive RT-PCR), nuclease protection assay (RNase, S1 nuclease assay), in situ hybridization, nucleic acid microarray, Northern blot, or a composition for diagnosis of liver cancer performed by a method selected from the group consisting of a DNA chip. The method according to claim 6, wherein the agent for measuring the expression level of the biomarker gene at the protein level is an antibody, antibody fragment, aptamer, or avidity that specifically recognizes the full-length protein of the marker or a fragment thereof. multimer) or peptidomimetic (peptidomimetics), a composition for diagnosing liver cancer. The method of claim 12, wherein the measurement is Western blot, ELISA (enzyme linked immunosorbent assay), radioimmunoassay (RIA), radioimmunodiffusion, immune electrophoresis, tissue immunostaining, immunoprecipitation assay (Immunoprecipitation assay) , Complement Fixation Assay (Complement Fixation Assay), FACS, mass spectrometry or a method selected from the group consisting of protein microarray, liver cancer diagnosis composition. A liver cancer diagnostic kit comprising the composition of claim 6 . A pharmaceutical composition for preventing or treating liver cancer comprising a SMARCA4 inhibitor as an active ingredient. The pharmaceutical composition for preventing or treating liver cancer according to claim 15, further comprising SMARCA2 or its expression promoter, SMARCC1 inhibitor, or IRAK1 inhibitor. The pharmaceutical composition for preventing or treating liver cancer according to claim 15, wherein the liver cancer is liver cancer in which IRAK1 is overexpressed. The pharmaceutical composition for preventing or treating liver cancer according to claim 15, wherein the liver cancer is hepatocellular carcinoma. (a) measuring the gene expression level of SMARCA4 or IRAK1 in a biological sample isolated from the test subject;
(b) comparing the corresponding gene of the normal control sample with the corresponding result; and
(c) when the gene expression level in step (a) is higher than the expression level of the corresponding gene in step (b), providing information necessary for diagnosing liver cancer, including determining that the test subject is liver cancer Way. 20. The method of claim 19,
(a) measuring the gene or protein expression level of SMARCC1, SMARCA2, Gankyrin, or AKR1B10 in a biological sample isolated from the test subject;
(b) comparing the corresponding gene or protein of the normal control sample with the corresponding result; and
(c) When the expression level of SMARCC1, Gankyrin or AKR1B10 in step (a) is high compared to the expression level of the corresponding gene or protein in step (b), or the expression level of SMARCA2 in step (a) in step (b) When the expression level of the corresponding gene or protein is low, the method of providing information necessary for diagnosing liver cancer, further comprising the step of determining that the test subject is liver cancer. (a) measuring the gene expression level of SMARCA4 or IRAK1 in a biological sample isolated from the test subject;
(b) comparing the corresponding gene of the normal control sample with the corresponding result; and
(c) when the gene expression level in step (a) is higher than the expression level of the corresponding gene in step (b), information for predicting the prognosis of liver cancer, including the step of determining that the prognosis of the test subject is bad How to provide. A method for predicting liver cancer treatment responsiveness of a SMARCA4 inhibitor, comprising the step of measuring the expression level of IRAK1. The method of claim 22, further comprising the step of measuring the expression level of Gankyrin or AKR1B10, predicting the responsiveness of the SMARCA4 inhibitor to the treatment of liver cancer.

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