KR20230088634A - Liver cancer specific biomarkers and use thereof - Google Patents

Abstract

The present invention relates to the use of genes whose expression changes specifically in hepatocellular carcinoma as biomarkers for detecting and diagnosing hepatocellular carcinoma and for treating hepatocellular carcinoma using these as targets. In the present invention, the expression of SMARCA4 is increased in HCC, It was revealed that SMARCA4 directly increases IRAK1 expression, and by confirming that IRAK1 induces oncoproteins Gankyrin and AKR1B10 through transcriptional activation, SMARCA4-IRAK1-Gankyrin and AKR1B10 can be used as liver cancer-specific markers, They can be usefully utilized 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 liver cancer-specific biomarkers and uses thereof, and more particularly, to use genes whose expression changes specifically in hepatocellular carcinoma as biomarkers for detecting and diagnosing hepatocellular carcinoma, and targeting them to treat liver cancer 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. The cause of cancer has not yet been identified, but it is considered that the complex factors of genetic factors, which are internal factors, carcinogenic chemicals that act as cancer-causing factors, continuous inflammation and damage, and cancer-causing viral infections are external factors. . However, cancer is not hopeless enough to be concluded as an incurable disease, and can be cured with early diagnosis and active treatment. Therefore, early detection, early diagnosis, and treatment are crucial to increasing the effectiveness of cancer treatment, and even advanced cancer can be cured or prolong life and improve painful symptoms if multifaceted and active methods are employed.

Among cancers, liver cancer is known as one of the most lethal cancers worldwide, and it is an aggressive cancer that ranks third in mortality (Ahn J, Flamm SL Hepatocellular carcinoma Dis Mon 2004;50:556-573). % 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 mechanisms of development and progression of liver cancer are still unknown (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 in 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 cause the clinical features observed in individual liver cancer patients. Liver cancer can be largely divided into primary liver cancer (hepatocellular carcinoma) that has occurred from the liver cells itself and metastatic liver cancer that has metastasized to the liver from cancer of other tissues. About 90% or more of liver cancer is primary liver cancer.

Hepatocellular carcinoma (HCC) is the fifth most common tumor in the world, accounting for 500,000 deaths each year (Okuda 2000). Survival rates for HCC patients have not improved over the past 20 years and have morbidity rates nearly equal to mortality rates (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 to be major risk factors for HCC (Thorgeirsson and Grisham 2002), and cell cycle mechanism Although it has been reported that changes in cell cycle regulators that progress from G1 to G1 are related to liver cancer formation (Hui et al, Hepatogasteroenterology 45:1635-1642, 1998), intracellular molecular mechanisms related to the onset and progression of liver cancer are unknown. It is still not clearly defined. According to conventional studies, protooncogenes such as various growth factor genes are mutated into oncogenes for various reasons and overexpressed or overactive, or tumors such as Rb protein or p53 protein When a tumor suppressor gene is mutated for various reasons and underexpressed or loses its function, it is reported to cause the onset and progression of various cancers, including liver cancer. In addition, DNA mutations and genetic alterations in gene expression have been reported to be 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 has been recognized that the onset and progression of most cancers, including liver cancer, is not caused by a few specific genes, but by complex interactions of various genes related to cell cycle and signal transduction. Therefore, individual genes or proteins The need for holistic research on various genes or proteins, away from focusing only on the expression or function of , is emerging.

An object of the present invention is 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 diagnosis 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.

In addition, an object of the present invention is to provide a method for providing information for predicting the prognosis of liver cancer.

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

To achieve the above object, the present invention provides a biomarker for diagnosing liver cancer comprising at least one gene selected from the group consisting of SMARCA4, SMARCC1, SMARCA2 and IRAK1 or a protein expressed from the gene.

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 for 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, and it was revealed that SMARCA4 directly increased IRAK1 expression, 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 liver cancer-specific markers, and they can be usefully utilized as targets for liver cancer treatment.

1 is a diagram confirming genetic variation and abnormal expression of SWI/SNF subunit genes in HCC:
A: mutation rate of SWI/SNF complex subunit genes 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 sets;
C: Differential gene expression of SMARCA4 , SMARCC1 and SMARCA2 in tissue pairs of HCC tissue and corresponding noncancerous liver biopsies of HCC;
D: Expression of SMARCA4 , SMARCC1 and SMARCA2 in tissue pairs of HCC tissues and corresponding noncancerous liver biopsies of HCC analyzed by qRT-PCR; and
E: Results of Western blot analysis of SMARCA4, SMARCC1 and SMARCA2 in 10 pairs of selected HCC subsets.
Figure 2 is a diagram analyzing the tumorigenicity (tumorigenicity) of SMARCA4, SMARCC1 and SMARCA2 in HCC:
A: cell viability of HCC treated with si-SMARCA4;
B: Cell proliferation rate of HCC treated with si-SMARCA4;
C: cell viability of HCC treated with si-SMARCC1;
D: Cell proliferation rate of HCC treated with si-SMARCC1; and
E: Cell viability of SMARCA4 high-expressing cells and SMARCA4 non-expressing cells treated with si-SMARCA2.
Figure 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: migrating cell image;
C: Cell cycle profile of si-SMARCA4 treated HCC cells;
D: result of Western blot analysis of cell cycle-related proteins in HCC cell lines;
E: si-Cont, si-Smarca4, si-Smarcc1 and pBJ-Smarca2 administration schedules, ultrasonography images, and tumor mass and mass numbers in the Ras -Tg mouse model; and
F: Results of Western blot analysis of Smarca4, Smarcc1 and Smarca2 in liver tissues of the Ras -Tg mouse model.
Figure 4 is a diagram confirming specific target genes regulated by SMARCA4 in HCC:
A: Pie charts, Venn diagrams and schematics of enhancer genes and SMARCA4-disrupted genes;
B: Heatmap of 563 SMARCA4-related genes in multistage HCC;
C: functional analysis of 563 genes;
D: Expression analysis results of SMARCA4 and IRAK1 in HCC cell lines treated with si-SMARCA4 (qRT-PCR);
E: Results of ChIP-qPCR analysis to evaluate SMARCA4 binding to the IRAK1 enhancer; and
F: Dual luciferase assay results in HCC cell lines expressing a reporter plasmid expressing the IRAK1 enhancer region after treatment with si-SMARCA4.
5 is a diagram confirming overexpression of SMARCA4 and IRAK1 in an 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 IRAK1 gene;
B: cell growth (cell viability) and cell proliferation rate after SMARCA4 or IRAK1 gene knockdown;
C: cell cycle profile;
D: result of Western blot analysis of cell cycle related proteins;
E: Cell images and cell numbers identified using Boyden chamber motility analysis; and
F: Cell images and cell numbers confirmed using the Transwell invasion assay.
Figure 7 is a diagram confirming the regulation of IRAK1 activity of SMARCA4 in HCC:
A: anti-tumor effect by IRAK1 (pcDNA3.1_IRAK1) after knocking down SMARCA4 (Western blot analysis, MTT analysis and BrdU analysis);
B: cell cycle profile;
C: result of cell cycle regulatory protein confirmed by Western blot analysis;
D: Cell images and cell numbers determined using Boyden chamber motility analysis; and
E: Cell images and cell numbers confirmed using Transwell invasion assay.
Figure 8 is a diagram confirming that SMARCA4 activates tumor proteins Gankyrin and AKR1B10 by regulating IRAK1 in HCC:
A: Western blot analysis of IRAK1 downstream regulatory molecules;
B: Western blot analysis of expression changes of IRAK1 downstream regulatory molecules by SMARCA4 knockdown (si-SMARCA4) and IRAK1 recovery (pcDNA3.1_IRAK1);
C: dosing schedule of the Ras -Tg mouse model;
D: tumor mass number;
E: Western blot analysis result of liver tissue of Ras -Tg mouse; and
F: Mechanistic diagram of SMARCA4-IRAK1-Gankyrin and AKR1B10.

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

In addition, the terms used in this specification (terminology) are terms used to appropriately express preferred embodiments of the present invention, which may vary according to the intention of a user or operator or customs in the field to which the present invention belongs. Therefore, definitions of these terms will have to be made based on the content throughout this specification. Throughout the specification, when a certain component is said to "include", it means that it may further include other components without excluding other components unless otherwise stated.

As used herein, the term "subject" or "patient" refers to any single individual in need of treatment, including humans, apes, monkeys, cows, dogs, guinea pigs, rabbits, chickens, insects, and the like. Also included are any subjects participating in clinical research trials who do not show any clinical signs of disease or subjects participating in epidemiological studies or subjects used as controls.

As used herein, the term "sample (sample)" means a biological sample obtained from a subject or patient. The source of the biological sample may be solid tissue from a fresh, frozen and/or preserved organ or tissue sample or biopsy or aspirate; blood or any blood component; It may be a cell at any point in the subject's pregnancy or development.

All technical terms used in the present invention, unless defined otherwise, are used with the same meaning as commonly understood by one of ordinary skill in the art related to the present invention. In addition, although preferred methods or samples are described in this specification, those similar or equivalent thereto are also included in the scope of the present invention. The contents of all publications incorporated herein 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 at least one gene or a protein expressed from the gene.

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

In one embodiment, the liver cancer may be hepatocellular carcinoma (HCC), more preferably hepatocellular carcinoma in which IRAK1 is overexpressed.

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

In one embodiment, the expression of SMARCA4, SMARCC1, IRAK1, Gankyrin or AKR1B10 may be increased specifically for liver cancer, and the expression of SMARCA2 may be decreased specifically for liver cancer.

In one aspect, the present invention relates to a composition for diagnosing liver cancer, including 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 include 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 in which IRAK1 is overexpressed.

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

In one embodiment, the agent for measuring the expression level of the biomarker gene at the mRNA level is a 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 complementary sequence. 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 (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 DNA chip.

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

As used herein, the term "detection" or "measurement" means detecting or quantifying the concentration of a measured object.

As used herein, the term "primer" is a nucleic acid sequence having a short free 3-terminal hydroxyl group, capable of forming a base pair with a complementary template, and serving as a starting point for template strand copying. It refers to a short nucleic acid sequence that functions. Primers can initiate DNA synthesis in the presence of reagents for polymerization (i.e., DNA polymerate or reverse transcriptase) and four different nucleoside triphosphates in an appropriate buffer and temperature.

In the present invention, the term "probe" refers to a nucleic acid fragment such as RNA or DNA corresponding to a few bases to several hundred bases in length that can form a specific binding with mRNA, and is labeled to confirm 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, or an RNA probe. In the present invention, hybridization is performed using probes complementary to the AFP, HMMR, NXPH4, PITX1, THBS4, and/or UBE2T genes, and the degree of expression of the gene can be diagnosed through hybridization. Selection of an appropriate probe and hybridization conditions may be modified based on those known in the art, so 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 can also be modified using a number of means known in the art. Non-limiting examples of such modifications include methylation, capping, substitution of one or more homologues of a natural nucleotide, and modifications between nucleotides, such as uncharged linkages such as methyl phosphonates, phossotriesters, phosphoro amidates, carbamates, etc.) or to 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 steps by an optimization procedure. This procedure is carried out as a series of procedures by those skilled in the art to establish a protocol for use in the laboratory. For example, conditions such as temperature, concentration of components, hybridization and washing time, 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, Nucleic Acid Hybridization, Springer-Verlag New York Inc. N.Y. (1999). For example, among the above stringent conditions, high stringency conditions are hybridization at 65 ° C in 0.5 M NaHPO 4 , 7% sodium dodecyl sulfate (SDS) and 1 mM EDTA, and 68 ° C in 0.1 x standard saline citrate (SSC) / 0.1% SDS. It means washing under conditions. Alternatively, high stringency means washing in 6 x SSC/0.05% sodium pyrophosphate at 48°C. Low stringency means, for example, washing at 42° C. in 0.2 x SSC/0.1% SDS.

As used herein, the term "antibody" is a term known in the art and refers to a specific protein molecule directed against an antigenic site. For the purpose of the present invention, an antibody refers to an antibody that specifically binds to a protein expressed in the AFP, HMMR, NXPH4, PITX1, THBS4 and/or UBE2T gene, which is a marker of the present invention, and the method for preparing the antibody is widely used. It can be prepared using a known method. This 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 polyclonal antibody, a monoclonal antibody, or any antibody having antigen-binding properties is also included in the antibody of the present invention, and all immunoglobulin antibodies are included. Furthermore, the antibodies of the present invention include special antibodies such as humanized antibodies.

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

In one embodiment, the kit may further comprise 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. there is. For example, it may include PCR primers to amplify relevant regions of genomic DNA. The kit may contain probes of genetic factors useful for pharmacogenomic profiling. Also, in the use of such kits, labeled oligonucleotides can be used for easy identification during analysis.

In the present invention, the term “diagnosis” as used herein refers to determining the susceptibility of a subject to a specific disease or disorder, determining whether a subject currently has a specific disease or disorder, determining whether a specific disease or disorder Determining the prognosis of a subject with a disease (e.g., identifying pre-metastatic or metastatic cancer status, determining the stage of a cancer, or determining the responsiveness of a cancer to treatment), or using therametrics (e.g., monitoring the condition of the subject to provide information on the efficacy of the treatment).

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

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, and may be siRNA of SEQ ID NO: 1.

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

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

In one embodiment, the liver cancer may be hepatocellular carcinoma, more preferably hepatocellular carcinoma in which IRAK1 is overexpressed.

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. In the above, "pharmaceutically acceptable" refers to a composition that is physiologically acceptable and does not cause an allergic reaction such as gastrointestinal disorder, dizziness, or similar reaction when administered to humans. Pharmaceutically acceptable carriers include, for example, carriers for oral administration such as lactose, starch, cellulose derivatives, magnesium stearate and stearic acid, and carriers for parenteral administration such as water, suitable oil, saline, aqueous glucose and glycol, etc. and the like, and may further include a stabilizer and a preservative. Suitable stabilizers include antioxidants such as sodium bisulfite, sodium sulfite or ascorbic acid. Suitable preservatives include 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 forms for parenteral or oral administration according to known methods, and a representative formulation for parenteral administration is an isotonic aqueous solution or suspension for injection. Formulations for injection may be prepared according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. For example, it can be formulated for injection by dissolving each component in saline or buffer. In addition, dosage forms for oral administration include, but are not limited to, powders, granules, tablets, pills, and capsules. The pharmaceutical composition formulated in the above way may be administered in an effective amount through various routes including oral, transdermal, subcutaneous, intravenous or intramuscular. It means that the administration route of the substance can be administered through any general route as long as it can reach the target tissue. In the above, "effective amount" refers to an amount that exhibits 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 cells, etc. can be easily determined by experts in In addition, the pharmaceutical composition of the present invention may be administered in combination with conventional therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple times. Preferably, considering all of the above factors, it is possible to administer an amount that can obtain the maximum effect with the minimum amount without side effects, more preferably 1 to 10000 μg/kg body weight/day, and even more preferably 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 may include, preferably, a chemical substance, a nucleotide, a vector containing the gene, a protein in which the gene is translated or a fragment thereof, or a natural product extract as an active ingredient.

As used herein, the term “stimulation of expression” means to cause enhancement of expression (into mRNA) or translation (into protein) of a target gene.

In one embodiment, the expression inhibitor of SMARCC1, IRAK1, Gankyrin or AKR1B10 may preferably include a chemical substance, nucleotide, antisense, siRNA oligonucleotide or natural product extract as an active ingredient, more preferably a 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 "expression inhibition" means to cause a reduction in the expression (into mRNA) or translation (into protein) of a target gene, preferably by which the expression of the target gene is undetectable or insignificant. means to exist.

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 /29058, 99/07409 and 00/44914). Since siRNA can suppress the expression of a target gene, it is provided as an efficient gene knock-down method or 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 each other, It may have a double-stranded structure, and also, 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 complete pairing of double-stranded RNA parts paired with each other, but pairing by mismatch (corresponding bases are not complementary), bulge (no base corresponding to one chain), etc. Unfinished parts may be included. In addition, the siRNA end structure can be either a blunt end or a cohesive end as long as it can suppress the expression of the SMARCC1, IRAK1, Gankyrin or AKR1B10 gene by the RNAi effect, and the cohesive end structure is a 3'-terminal protruding 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, in which case the siRNA molecule formed by expression of the nucleotide sequence is resistant to intramolecular hybridization. By this, a hairpin structure is formed, and a stem-and-loop structure is formed as a whole (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 producing siRNA include direct synthesis of siRNA in a test tube and introduction into cells through a transformation process, and transfection of siRNA expression vectors or PCR-derived siRNA expression cassettes prepared so that siRNA is expressed in cells into cells. There is a way to convert or infect.

In one embodiment, the composition of the present invention including the gene-specific siRNA may include an agent that promotes the entry of siRNA into cells. Agents that promote siRNA uptake into cells can generally use agents that promote nucleic acid uptake. For example, liposomes are used or one of a number of sterols including cholesterol, cholate, and deoxycholic acid is used. It can be formulated with an oily carrier. In addition, poly-L-lysine, spermine, polysilazane, polyethylenimine (PEI), polydihydroimidazolenium, polyallylamine ), a cationic polymer such as chitosan may be used, and succinylated PLL (succinylated PLL), succinylated PEI (polyglutamic acid), polyaspartic acid Anionic polymers such as polyaspartic acid, polyacrylic acid, polymethacylic acid, dextran sulfate, heparin, and hyaluronic acid available.

In one embodiment, when an antibody specific to the protein is used as a substance that reduces the expression and activity of the SMARCC1, IRAK1, Gankyrin or AKR1B10 protein, the antibody is directly coupled to an existing therapeutic agent or indirectly through a linker or the like (e.g. covalent bond).

As used herein, the term "antisense oligonucleotide" refers to DNA or RNA or derivatives thereof containing a nucleic acid sequence complementary to a specific mRNA sequence, and binds to a complementary sequence in mRNA to SMARCC1, IRAK1, Gankyrin or inhibits the translation of AKR1B10 into a protein The antisense sequence of the present invention refers to a DNA or RNA sequence that is complementary to SMARCC1, IRAK1, Gankyrin or AKR1B10 mRNA and capable of binding to SMARCC1, IRAK1, Gankyrin or AKR1B10 mRNA. In addition, the antisense nucleic acid can inhibit translation, cytoplasmic translocation, maturation, or any other essential activity for overall biological function of SMARCC1, IRAK1, Gankyrin or AKR1B10 mRNA. (De Mesmaeker et al., Curr Opin Struct Biol., 5, 3, 343-55, 1995). ates, phosphotriesters, methyl phosphonates, short-chain alkyls, cycloalkyls, short-chain heteroatomic, heterocyclic intersugar linkages, etc. In addition, antisense nucleic acids can contain one or more substituted sugar moieties. Antisense nucleic acids may include modified bases, including hypoxanthine, 6-methyladenine, 5-methylpyrimidine (particularly 5-methylcytosine), 5-hydroxymethylcytosine. (HMC), Glycosyl HMC, Gentobiocil HMC, 2-aminoadenine, 2-thiouracil, 2-thiothymine, 5-bromouracil, 5-hydroxymethyluracil, 8-azaguanine, 7-decane azaguanine, N6 (6-aminohexyl) adenine, 2,6-diaminopurine, etc. In addition, the antisense nucleic acid of the present invention may include one or more moieties that enhance the activity and cell adhesion of the antisense nucleic acid; It can be chemically combined with a conjugate. Cholesterol moiety, cholesteryl moiety, cholic acid, thioether, thiocholesterol, fatty chain, phospholipid, polyamine, polyethylene glycol chain, adamantane acetic acid, palmityl moiety, octadecylamine, hexylaminocarbonyl-oxycol and fat-soluble moieties such as esterol moieties, but are not limited thereto. Oligonucleotides containing fat-soluble moieties and preparation methods are already well known in the art (U.S. Patent Nos. 5,138,045, 5,218,105 and 5,459,255). The modified nucleic acid may increase stability to nucleases and increase binding affinity between the antisense nucleic acid and the target mRNA. The antisense oligonucleotide may be synthesized in vitro by a conventional method and administered into a living body, or the antisense oligonucleotide may be synthesized in vivo. One 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 allow antisense RNA to be transcribed using a vector in which the origin of the recognition site (MCS) is reversed. Such antisense RNA preferably has a translational stop codon in the sequence so that it is not translated into the peptide sequence.

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

In one embodiment, the method comprises (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 with corresponding results of the corresponding gene or protein in a normal control sample; And (c) when the expression level of SMARCC1, Gankyrin or AKR1B10 in step (a) is higher than the expression level of the corresponding gene or protein in step (b), or the expression level of SMARCA2 in step (a) (b) When lower than the expression level of the corresponding gene or protein in the step, a step of determining that the test subject has liver cancer may be further included.

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

In one aspect, the present invention comprises (a) measuring the gene expression level of SMARCA4 or IRAK1 in a biological sample isolated from a test subject; (b) comparing with the corresponding result of the gene of interest in a normal control sample; And (c) if the gene expression level in step (a) is higher than the expression level of the gene in step (b), for predicting the prognosis of liver cancer, including the step of determining that the test subject has a poor prognosis It is about how to present 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 include 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 for specifying the content of the present invention, and the present invention is not limited thereto.

Example 1. Evaluation of subunit gene expression levels of the 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). Level 3 mRNA expression data of TCGA-LIHC HTSeq-FPKM was log2 transformed and [log2(fpkm+1)] was used to evaluate gene expression levels. As a result, human HCC (cBioPortal; n=1,019) showed 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 dominant aberrant expression of a particular subunit gene can cause systemic disturbances in the SWI/SNF complex. Accordingly, as a result of examining the expression of SWI/SNF subunit genes using multilevel 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 at the same time, it was confirmed that SMARCA2 was downregulated (≥ ± 1.5 fold, P < 0.05). In addition, 34 HCC tissues and corresponding non-cancerous liver biopsy HCC tissue pairs were obtained from the National Biobank in Korea and analyzed. As a result, abnormal regulation of SMARCA4, SMARCA2 and SMARCC1 was consistent with TCGA_LIHC, ICGC_LIRI and GSE77314. Tissue pairs of HCC were also identified (Fig. 1C). In addition, further validation by qRT-PCR in an additional set of 34 randomly selected HCC tissues showed that SMARCA4 (67.6%) and SMARCC1 (52.9%) overexpression and SMARCA2 (61.8%) inhibition were consistently observed in this set ( Fig. 1D). In addition, as a result of analyzing 10 tissue pairs of HCC and non-cancerous livers selected by Western blot, significant overexpression of SMARCA4 and SMARCC1 was simultaneously suppressed by SMARCA2 in HCC (FIG. 1E).

Example 2. Antitumor effect by modulating the expression of SMARCA4, SMARCA2 and SMARCC1

2-1. in vitro analysis

In order to confirm the role of SMARCA4, SMARCA2 and SMARCC1, whose expression was specifically changed in HCC 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, which are hepatic cell lines with relatively high SMARCA4 and SMARCC1 expression and relatively low SMARCA2 expression than normal hepatocytes, and cell viability and cell proliferation were measured by MTT assay and BrdU. It was confirmed by analysis and clonogenic assay analysis. In addition, migration and invasion of HCC cells knocked down 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 regulation proteins were confirmed by Western blot analysis.

As a result, it was shown that the knockdown of SMARCA4 and SMARCC1 significantly suppressed the cell growth and proliferation rate of HCC cells (Fig. 2A to D). In addition, in both SMARCA4-null HCC cell lines, PLC/PRF/5 and SK-Hep-1, knockdown of SMARCA2 resulted in overall lethality of HCC cells. appeared to be absent (Fig. 2E). These results indicate a pivotal role for the SMARCA4-SWI/SNF complex in hepatocellular carcinogenesis. In addition, the Boyden chamber motility assay results showed that SMARCA4 knockdown significantly inhibited the serum-stimulated migration and invasion responses of HCC cells (FIG. 3A), and the wound healing assay results similarly showed that HCC cell wound healing It was shown that SMARCA4 knockdown reduced the effect (FIG. 3B). In addition, as a result of confirming the effect of SMARCA4 knockdown on cell cycle regulation of HCC cells by flow cytometry, it was found that SMARCA4 knockdown increased the number of cells in the G1 phase in HCC cells (Fig. 3C), and Western blotting showed that cell cycle As a result of confirming the expression of regulatory proteins, knockdown of SMARCA4 selectively induced p27 Kip1 expression in HCC cells, and at the same time inhibited cyclin D1, cyclin E, cyclin-dependent kinase 2 (CDK2), CDK4 and CDK6, resulting in pRb hyperphosphorylation (hypophosphorylation) (p-pRb) (Fig. 3D). Through this, it was found that the hyperactivity of SMARCA4 in HCC promotes the transition from G1 phase to S phase by regulating electrons of cell cycle proteins.

2-2. in vivo assay

To determine if modulation of SMARCA4, SMARCA2, and SMARCC1 results in tumor suppressive effects in vivo, eight Ras-Tg (H-ras12V homozygous transgenic) male mice in which HCC develops at approximately 15 to 18 weeks of age were examined for SMARCA4 from 14 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 when livers were obtained, and the weight and tumor mass number of the obtained livers were confirmed, and they were disrupted to analyze SMARCA4, SMARCA2 and SMARCC1 by Western blot analysis. The expression of was confirmed.

As a result, HCC was detected from 18 weeks of age in the control mice (si-Cont), and many large HCCs occurred in 3 out of 4 mice. On the other hand, no HCC was detected in the si-Smarca4-treated mice until 24 weeks of age. However, HCC was detected at 20 weeks of age in the si-Smarcc1-treated mice and the pBJ-Smarca2-treated mice, and relatively small HCC developed in 3 out of 4 mice. Total liver weight change and tumor mass number 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 liver tissues of each group of mice (FIG. 3F).

Example 3. Identification of SMARCA4-regulated genes in HCC

3-1. SMARCA4-regulated gene selection

To identify the SMARCA4-SWI/SNF regulated genes, by SMARCA4 in HCC cells using histone ChIP-seq of HepG2 cells available from Encyclopedia of DNA Elements (ENCODE) (https://www.encodeproject.org/). Positions enriched in the specifically recognized markers H3K27ac and H3K27me3 were comprehensively mapped. Here, the trimmed reads were aligned to the human genome using the Bowtie 2 ( http://bowtie-bio.sourceforge.net/bowtie2/index.shtml ) mapper, and to remove PCR bias, redundant reads The values were removed using the Sambamba tool ( https://lomereiter.github.io/sambamba/ ). HOMER (findPeaks) was used to identify positions enriched in H3K27ac or H3K27me3 (FDR-adjusted p-value cutoff=0.001). Through this analysis, 9,198 genes out of 21,684 genes closest to the peak were identified as active enhancers (FIG. 4A). Then, in order to identify gene elements regulated in HCC cells, SMARCA4 knockdown was performed in Hep3B to find 1,865 gene elements regulated by SMARCA4 (> ± 1.5 fold, P < 0.05), among which 1,054 genes were found to be downregulated in HCC cells in which SMARCA4 was knocked down. As a result of analyzing these 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-related genes showed gradual increase or decrease in transcript 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 in the 563 gene sets (Fig. 4C). ). To select genes regulated by SMARCA4 in HCC, we reduced 563 genes 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 involved, the top 5 genes upregulated were examined 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. Confirmation of interaction of SMARCA4 and IRAK1

In order 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 active enhancer regions of CKAP4 and IRAK1 (H3K27ac) enrichment rates were confirmed. Chromatin immunoprecipitation (ChIP) analysis was performed according to the instructions of the manufacturer (Pierce Agarose ChIP kit; Thermo Fisher Scientific, Waltham, MA), and RT-RT-DNA was prepared using primers for the enhancer region of each candidate gene regulated by SMARCA4. Amplified by qPCR. Cross-linked DNA was precipitated using SMARCA4 antibody (anti-SMARCA4) and acetylated histone H3 at lysine 27 (Abcam, Cambridge, UK) antibody (anti-H3K27ac). The experiment was repeated three times and averaged normalized to IgG. In addition, to confirm that endogenous SMARCA4 directly binds to the IRAK1 enhancer region, the IRAK1 enhancer was cloned into the pGL4.23 reporter vector, followed by a dual luciferase reporter assay of the pGL4.23_IRAK1 enhancer in the presence or absence of SMARCA4 in HCC cells. performed.

As a result, the enrichment rate of the IRAK1 active 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 active enhancer in the same cells The region was consistently unaffected by SMARCA4 knockdown (Fig. 4E). In addition, as a result of the luciferase reporter assay, SMARCA4 knockdown significantly suppressed the relative luciferase activity in all HCC cells, confirming the interaction between SMARCA4 and the IRAK1 enhancer (FIG. 4F).

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

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

Example 5. Confirmation 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, changes in IRAK1 expression were confirmed in TCGA_LIHC and ICGC_LIRI. As a result, SMARCA4 of IRAK1 A strong positive correlation with was observed (Fig. 6A). In addition, Kaplan-Meier survival curves 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). Accordingly, 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 flow cytometry analysis of PI-stained cells to confirm cell cycle regulation, knockdown of SMARCA4 and IRAK1, respectively, induced cell cycle arrest in 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, knockdown of SMARCA4 and IRAK1 reduced the migration and invasion responses of chemoattractant-stimulated HCC cells (FIGS. 6E and F).

Example 6. Regulation of IRAK1 activity by SMARCA4

To confirm that SMARCA4 directly regulates IRAK1 activity, HCC cells were knocked down with SMARCA4 siRNA, and recovery experiments were 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 these anti-tumor effects were abolished by co-transformation of the IRAK1 plasmid (FIG. 7A). In addition, as a result of flow cytometric analysis of PI-stained cells, knockdown of SMARCA4 in HCC cells markedly induced G1-phase arrest and exhibited an anti-growth effect, whereas co-transformation of the IRAK1 plasmid in the same cells resulted in a reduction in the HCC cell cycle. It was not stopped and tumor growth resumed again (Fig. 7B). These results were further verified by identifying cell cycle-related proteins by Western blot analysis. As a result, p27 Kip1, which was selectively induced by SMARCA4 knockdown in HCC, was suppressed by IRAK1 plasmid expression, and simultaneously by SMARCA4 knockdown. Phosphorylation of cyclin D1, cyclin E, CDK2, CDK4, and CDK6 whose expression was suppressed, and pRb was restored (FIG. 7C). In addition, chemoattractant-stimulated migration and invasion responses reduced by SMARCA4 knockdown in HCC cells were also restored due to expression of the IRAK1 plasmid (FIGS. 7D and E).

Example 7. Oncoprotein regulation of IRAK1

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

<110> NEORNAT <120> LIVER CANCER SPECIFIC BIOMARKERS AND USE THEREOF <130> PN2305-238 <150> KR 10-2020-0090126 <151> 2020-07-21 <160> 2 <170> KoPatentIn 3.0 <210> 1 <211> 19 <212> RNA <213> artificial sequence <220> <223> si-SMARCA4 <400> 1 cucucucaac gcuguccaa 19 <210> 2 <211> 25 <212> RNA <213> artificial sequence <220> <223> si-IRAK1 <400> 2 agaacggcuu cuacugccug gugua 25

Claims (4)

A pharmaceutical composition for preventing or treating liver cancer in patients with increased expression of IRAK1, Gankyrin or AKR1B10, comprising siRNA represented by the nucleotide sequence of SEQ ID NO: 1 as an active ingredient. According to claim 1,
The liver cancer is hepatocellular carcinoma (HCC), a pharmaceutical composition for preventing or treating liver cancer According to claim 1,
A pharmaceutical composition for preventing or treating liver cancer, characterized in that the siRNA represented by the nucleotide sequence of SEQ ID NO: 1 inhibits the expression of SMARCA4 According to claim 1,
Characterized in that the liver cancer is liver cancer of a patient with increased expression of IRAK1, Gankyrin and AKR1B10, a pharmaceutical composition for preventing or treating liver cancer

Images