JP2011103821A - Identification of gene with variable expression, responding by stimulation of sorafenib - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for predicting the response to the stimulation of agents by the use of genetic polymorphism and to provide markers used in the method for predicting the response to the stimulation of the agents. <P>SOLUTION: The method includes the prediction of the response to Sorafenib of samples administered with Sorafenib by detecting in vitro genetic polymorphisms which decide the difference in the amount of expression of any one of the following (a)-(h) genes which show the difference between alleles in the amount of expression in response to the stimulation of Sorafenib: (a) dihydropyrimidine dehydrogenase gene, (b) CD247 molecule gene, (c) phospholipase A2, group IVA (cytosolic, calcium-dependent) gene, (d) interleukin 1 receptor, typeII gene, (e) dedicator of cytokinesis 2 gene, (f) estrogen receptor 1 gene, (g) complement component 5 gene, (h) phospholipase C, beta 1 (phosphoinositide-specific) gene, (i) pancreatic lipase-related protein 3 gene, and (j) lipase, hepatic gene. <P>COPYRIGHT: (C)2011,JPO&INPIT

Description

  The present invention relates to identification of genes having expression level variability in response to Sorafenib stimulation and polymorphisms of the genes.

  The phenomenon in which even if the same gene is located at the same position in the genome and is on a different allele, the difference in the gene expression level is a relatively new concept that has been reported in recent years (Non-patent Document 1). See).

  There are two types of genes that show different expression among alleles: genes that are imprinted (imprinted genes) and genes that are not imprinted (non-imprinted genes). The former is called imprinting. When a single allele is inherited from a parent in one cell or tissue, it is inactivated by expression of either allele being physiologically modified such as methylation. This phenomenon is suppressed. Even in the latter case, that is, in a non-imprinted gene, different expression differences may be observed between alleles. This is because genomic polymorphism within or between alleles within a gene acts as a cis-acting element that regulates the expression of nearby genes, creating differences in gene expression between alleles. It is believed that. Changes in the expression of each allele due to differences in the genomic DNA sequence seen in the latter are considered to be inherited from generation to generation, resulting in differences in gene expression levels among individuals, as well as differences in individual constitutions, pathological conditions. And its risk, and the difference in drug responsiveness. Therefore, genes with different expression levels between alleles may be related to phenotypes such as diseases or disorders.

  On the other hand, the diversity of gene expression in response to drugs is that TNF / LTA shows allele-specific expression under conditions of 4-β-phorbol-12-myristate-13-acetate (PMA) + ionomycin stimulation. Has been reported (see Non-Patent Documents 2 and 3).

  As described above, the presence of diversity in the expression level of genes that respond to drugs is considered to be applicable to pharmacogenomics (PGx) studies. With PGx research, we look for genetic polymorphisms (SNP: Single Nucleotide Polymorphisms) that affect the dynamics, effects, and side effects of drugs from genetic analysis. The purpose is to stratify the patients shown. There are two methods of PGx research: Unbiased genome-wide approach that measures genomic DNA as comprehensively as possible without selecting candidates, or Candidate gene approach that selects and verifies candidates, including SNP maps, haplotype markers, The expression of genes involved in pharmacological functions and therapeutic effects, or the diversity among individuals in mutation in inactivation has been investigated. Therefore, in PGx research in drug discovery, an attempt is made to stratify patients who show favorable reactions to drugs and patients who show undesirable reactions. However, the Unbiased genome-wide approach requires a large number of samples and high costs, and the Candidate gene approach requires selection of candidates from a large number of gene polymorphisms that may be involved in drug action.

  Therefore, recently, a new method has been considered in which genomes are comprehensively measured in advance in advance, and candidate genes are selected by combining both approaches. Specifically, it is a method of exhaustively measuring genomic DNA and RNA expression ex vivo, and searching for SNPs that affect RNA expression fluctuations by drugs. All of the reported examples so far have investigated the relationship between mRNA expression fluctuations caused by drugs and SNP, and searched for SNPs that affect mRNA expression, which is called the Expression quantitative trait loci (eQTL) method (non-patented) (Ref. 4 and 5).

  On the other hand, the ExpressGenotyping method (see Patent Document 1) can comprehensively measure the expression level and total amount of all premature RNAs that have not undergone splicing at the same time as genomic DNA SNP typing, so only qualitative allele frequency data is available. Compared to, the reliability is high, and effective SNPs that affect the expression can be searched even with a small number of samples. Therefore, if this ExpressGenotyping method can be used to identify genes with diverse expression levels of drug responsiveness, drug side effects can be avoided in drug discovery, and patients with effective responses and patients with ineffective responses can be divided into layers. It is thought that it can be separated.

Japanese Patent No.4111985

Knight JC., Trends Genet. Mar; 20 (3): 113-6. PMID: 15049300, 2004 Knight JC. Et al., Nat Genet. Apr; 33 (4): 469-75. PMID: 12627232, 2003 Knight JC. Et al, Nat Genet. Apr; 36 (4): 394-9. PMID: 15052269, 2004 Cheung VG. Et al., Nature. Oct 27; 437 (7063): 1365-9. PMID: 16251966, 2005 Huang RS. Et al., Proc Natl Acad Sci U S A. Jun 5; 104 (23): 9758-63. PMID: 17537913, 2007

  An object of the present invention is to identify genes having a variety of expression levels of drug responsiveness in drug stimulation using ExpressGenotyping, which can determine genes whose expression levels differ between alleles. It is another object of the present invention to provide a method for searching for a gene polymorphism involved in a phenotype of a drug stimulus response by the method. Another object of the present invention is to provide a method for predicting a drug stimulus response using the gene polymorphism thus searched and a marker for detecting the drug polymorphism used in the method.

  As a result of intensive studies to solve the above problems, the present inventor believes that by using the anticancer agent Sorafenib as a drug, a gene whose expression level is different between alleles can be found efficiently. Actually, when ExpressGenotyping was performed after sorafenib stimulation, SNPs that can determine genes with different expression levels between alleles were successfully identified, and the present invention was completed. Sorafenib is used as a multikinase inhibitor in the treatment of kidney cancer, and as side effects, hypertension, skin disorders, diarrhea, and pain are known. In particular, skin disorders that occur frequently in Japanese are a major obstacle to treatment.

That is, the present invention is as follows.
[1] In vitro detection of a gene polymorphism that can determine the difference in expression level in any of the following genes (a) to (j), in which the expression level varies between alleles due to sorafenib stimulation Methods for predicting responsiveness of sorafenib in a subject receiving sorafenib:
(a) the dihydropyrimidine dehydrogenase (DPYD) gene;
(b) the CD247 molecule gene;
(c) phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene;
(d) interleukin 1 receptor, type II (IL1R2) gene;
(e) dedicator of cytokinesis 2 (DOCK2) gene;
(f) estrogen receptor 1 (ESR1) gene;
(g) complement component 5 (C5) gene;
(h) phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene;
(i) the pancreatic lipase-related protein 3 (PNLIPRP3) gene; and
(j) lipase, hepatic (LIPC) gene.

[2] The method for predicting responsiveness of sorafenib in a subject administered with sorafenib according to [1], wherein the gene polymorphism in which the difference in expression level in (a) to (j) can be determined is any of the following:
(i) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any one of SEQ ID NOs: 1 to 8 in the dihydropyrimidine dehydrogenase (DPYD) gene;
(ii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 9 in the CD247 molecule gene;
(iii) A single nucleotide polymorphism present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 10 to 12 in the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene Site;
(iv) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 13 to 15 in the interleukin 1 receptor, type II (IL1R2) gene;
(v) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 16 to 23 in the dedicator of cytokinesis 2 (DOCK2) gene;
(vi) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 24 or 25 in the estrogen receptor 1 (ESR1) gene;
(vii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 26 in the complement component 5 (C5) gene;
(viii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 27 to 46 in the phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene;
(ix) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 47 in the pancreatic lipase-related protein 3 (PNLIPRP3) gene; or
(x) A single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 48 to 52 in the lipase, hepatic (LIPC) gene.

[3] A method for predicting responsiveness of sorafenib in a subject administered with sorafenib according to [1], wherein the gene polymorphism in which the difference in expression level in (a) to (j) can be determined is any of the following:
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, the single nucleotide polymorphism rs1431485, the gene polymorphism present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of
(x) single nucleotide polymorphisms in lipase, hepatic (LIPC) genes rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542, and gene polymorphisms in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in

[4] Sorafenib responsiveness prediction is a prediction of the effect of sorafenib in a subject or a prediction of the type or extent of side effects of the subject's sorafenib administered with any of the sorafenibs of [1]-[3] A method of predicting responsiveness.

[5] An oligonucleotide consisting of a DNA fragment containing the following single nucleotide polymorphic site of (i) to (x): from 10 to 30 bases of the following genes (i) to (x) A probe for predicting responsiveness of sorafenib in a subject to which sorafenib is administered, comprising an oligonucleotide consisting of a partial sequence or an oligonucleotide complementary to the partial sequence or a label thereof:
(i) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 1 to 8 in the dihydropyrimidine dehydrogenase (DPYD) gene;
(ii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 9 in the CD247 molecule gene;
(iii) A single nucleotide polymorphism present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 10 to 12 in the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene Site;
(iv) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 13 to 15 in the interleukin 1 receptor, type II (IL1R2) gene;
(v) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 16 to 23 in the dedicator of cytokinesis 2 (DOCK2) gene;
(vi) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 24 or 25 in the estrogen receptor 1 (ESR1) gene;
(vii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 26 in the complement component 5 (C5) gene;
(viii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 27 to 46 in the phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene;
(ix) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 47 in the pancreatic lipase-related protein 3 (PNLIPRP3) gene; or
(x) A single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 48 to 52 in the lipase, hepatic (LIPC) gene.

[6] Any of the following oligonucleotides comprising a DNA fragment containing a single nucleotide polymorphic site of (i) to (x), comprising 10 to 30 bases of the following genes (i) to (x) A probe for predicting responsiveness of sorafenib in a subject to which sorafenib is administered, comprising an oligonucleotide consisting of a partial sequence or an oligonucleotide complementary to the partial sequence or a label thereof:
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, a single nucleotide polymorphism rs1431485, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of types, or
(x) single nucleotide polymorphisms in lipase, hepatic (LIPC) genes rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542, and gene polymorphisms in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in

[7] Sorafenib responsiveness prediction is a prediction of the effect of sorafenib in a subject or a prediction of the type or extent of side effects, wherein the responsiveness of sorafenib in a subject administered with [5] or [6] sorafenib Probe for prediction.

[8] An oligonucleotide consisting of a DNA fragment containing the following single nucleotide polymorphic sites (i) to (x): a partial sequence consisting of 10 to 30 bases of the following genes (i) to (x) Alternatively, an immobilized substrate for predicting responsiveness of sorafenib in a subject to which sorafenib is administered, to which at least one oligonucleotide having a sequence complementary to the partial sequence or a label thereof is immobilized:
(i) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 1 to 8 in the dihydropyrimidine dehydrogenase (DPYD) gene;
(ii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 9 in the CD247 molecule gene;
(iii) A single nucleotide polymorphism present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 10 to 12 in the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene Site;
(iv) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 13 to 15 in the interleukin 1 receptor, type II (IL1R2) gene;
(v) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 16 to 23 in the dedicator of cytokinesis 2 (DOCK2) gene;
(vi) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 24 or 25 in the estrogen receptor 1 (ESR1) gene;
(vii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 26 in the complement component 5 (C5) gene;
(viii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 27 to 46 in the phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene;
(ix) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 47 in the pancreatic lipase-related protein 3 (PNLIPRP3) gene; or
(x) A single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 48 to 52 in the lipase, hepatic (LIPC) gene.

[9] An oligonucleotide consisting of a DNA fragment containing the following single nucleotide polymorphic sites (i) to (x): a partial sequence consisting of 10 to 30 bases of the following genes (i) to (x) Alternatively, an immobilized substrate for predicting responsiveness of sorafenib in a subject to which sorafenib is administered, to which at least one oligonucleotide having a sequence complementary to the partial sequence or a label thereof is immobilized:
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, a single nucleotide polymorphism rs1431485, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of types, or
(x) single nucleotide polymorphisms in lipase, hepatic (LIPC) genes rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542, and gene polymorphisms in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in

[10] The prediction of sorafenib responsiveness in a subject who is administered sorafenib according to [8] or [9], wherein the prediction of responsiveness of sorafenib is a prediction of the effect of sorafenib in a subject or a prediction of the type or degree of side effects. Immobilization substrate for prediction.

[11] At least a pair of primer sets used for amplification of DNA fragments containing the following single nucleotide polymorphic sites (i) to (x), wherein the DNA polymorphisms of the following genes (i) to (x) A test administered with sorafenib consisting of an oligonucleotide consisting of a partial sequence consisting of 10 to 30 bases present on the 3 ′ side and 5 ′ side of at least one polymorphic site, or a sequence complementary to the partial sequence. A pair of primer sets to predict sorafenib responsiveness in the body:
(i) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 1 to 8 in the dihydropyrimidine dehydrogenase (DPYD) gene;
(ii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 9 in the CD247 molecule gene;
(iii) A single nucleotide polymorphism present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 10 to 12 in the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene Site;
(iv) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 13 to 15 in the interleukin 1 receptor, type II (IL1R2) gene;
(v) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 16 to 23 in the dedicator of cytokinesis 2 (DOCK2) gene;
(vi) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 24 or 25 in the estrogen receptor 1 (ESR1) gene;
(vii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 26 in the complement component 5 (C5) gene;
(viii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 27 to 46 in the phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene;
(ix) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 47 in the pancreatic lipase-related protein 3 (PNLIPRP3) gene; or
(x) A single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 48 to 52 in the lipase, hepatic (LIPC) gene.

[12] The following (i) to (x) at least a pair of primer sets used for amplification of a DNA fragment containing a single nucleotide polymorphic site, wherein the DNA polymorphisms of the following genes (i) to (x) A test administered with sorafenib consisting of an oligonucleotide consisting of a partial sequence consisting of 10 to 30 bases present on the 3 ′ side and 5 ′ side of at least one polymorphic site, or a sequence complementary to the partial sequence. A pair of primer sets to predict sorafenib responsiveness in the body:
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, a single nucleotide polymorphism rs1431485, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of types, or
(x) single nucleotide polymorphisms in lipase, hepatic (LIPC) genes rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542, and gene polymorphisms in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in

[13] Sorafenib responsiveness prediction is a prediction of the effect of sorafenib in a subject or a prediction of the type or extent of side effects, wherein the responsiveness of sorafenib in a subject administered with [11] or [12] sorafenib A pair of primer sets for prediction.
[14] Sorafenib comprising the probe of any one of [5] to [7], the immobilized substrate of any of [8] to [10], or the pair of primer sets of any of [11] to [13] A kit for predicting responsiveness of sorafenib in a subject to be administered.
[15] To determine the responsiveness of sorafenib in a subject administered with sorafenib according to [14], wherein the prediction of responsiveness of sorafenib is a prediction of the effect of sorafenib or a prediction of the type or degree of side effects in the subject kit.

  According to the present invention, it is possible to quickly and efficiently search for gene polymorphisms that determine genes whose expression levels differ between alleles in Sorafenib stimulation. Moreover, since the gene polymorphism searched in this way can identify the expression level between alleles, it can be an effective means for analyzing the phenotype involving the gene. Furthermore, by detecting the genetic polymorphism so searched, predict and evaluate the phenotype of the subject to be treated with sorafenib (main drug action or risk of developing side effects), In the future drug discovery in general, side effects of the drug can be avoided, and patients showing an effective response and patients showing an ineffective response can be stratified.

It is a figure which shows the result of the mRNA expression analysis analyzed by Human Genome U133 Plus 2.0 Array It is a figure which shows the result of the frequency distribution of Chromosome No. 1 in DMSO group of GM18940 cells. It is a figure which shows the gene polymorphism different among races which influence sorafenib stimulation. It is a figure which shows the 10 types of gene (gene polymorphism) EAI list | wrist which has a difference in expression level among alleles relevant to the side effect of sorafenib stimulation. It is a figure which shows the 10 types of gene (gene polymorphism) EAI list | wrist which has a difference in expression level among the alleles relevant to the side effect of sorafenib stimulation (continuation of FIG. 4-1). It is a figure which shows the position of the polymorphism in the base sequence of a dihydropyrimidine dehydrogenase (DPYD) gene. It is a figure which shows the position of the polymorphism in the base sequence of CD247 molecule gene. It is a figure which shows the position of the polymorphism in the base sequence of phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene. It is a figure which shows the position of the polymorphism in the base sequence of interleukin 1 receptor, type II (IL1R2) gene. It is a figure which shows the position of the polymorphism in the base sequence of dedicator of cytokinesis 2 (DOCK2) gene. It is a figure which shows the position of the polymorphism in the base sequence of estrogen receptor 1 (ESR1) gene. It is a figure which shows the position of the polymorphism in the base sequence of complement component 5 (C5) gene. It is a figure which shows the position of the polymorphism in the base sequence of phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene. It is a figure which shows the position of the polymorphism in the base sequence of pancreatic lipase-related protein 3 (PNLIPRP3) gene. It is a figure which shows the position of the polymorphism in the base sequence of a lipase, hepatic (LIPC) gene.

Hereinafter, the present invention will be described in detail.
1. BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a search for a gene whose expression level is different between alleles in response to sorafenib stimulation. Here, “a gene whose expression level is different between alleles” refers to a gene whose gene expression level from one allele is different from that of the other allele. The gene expression from each allele can be identified using a specific gene polymorphism as an index, but not all gene polymorphisms can distinguish the difference in the expression level between alleles. Moreover, it has not been known that a difference occurs in the expression level between alleles by stimulation with sorafenib. In the present invention, by using ExpressGenotyping method, the expression level differs between alleles against sorafenib stimulation by quickly and efficiently searching for gene polymorphisms that can determine the difference in expression level between alleles by sorafenib stimulation. A gene polymorphism (single nucleotide polymorphism; SNP) capable of discriminating a difference in gene expression between alleles can be searched for by searching for a gene.

  In this search, for example, the cultured cells were stimulated with sorafenib in an in vitro system and stimulated with a substance other than sorafenib, and gene expression in the cultured cells was comprehensively analyzed. May be selected, and a gene related to the difference in expression level between alleles of the gene polymorphism may be selected.

These methods can be performed by ExpressGenotyping method (the method described in Japanese Patent No. 4111985).
Hereinafter, typical techniques for searching for and selecting genes and gene polymorphisms will be described.

Cell culture Cell culture is a method known in the art. For example, a cell line is seeded in a cell culture medium (medium RPMI-1640 + 15% serum FBS), and cells are cultured at 37 ° C. and 5% CO ₂ . Culture can be performed.

Drug Stimulation Drug stimulation is a method known in the art. For example, two groups of a control group (DMSO) and a drug (sorafenib) can be added to a culture medium seeded with cultured cells to perform drug stimulation. .

RNA extraction Total RNA extraction is a method known in the art. For example, when total RNA is extracted, AGPC (acid guanidinium, phenol, chloroform method, etc.) or a commercially available kit can be used. .

mRNA expression
As a specific example of detecting the expression level of mRNA using a DNA chip, there is a method using GeneChip (registered trademark) Human Genome U133 Plus 2.0 Array manufactured by Affymetrix. The GeneChip (registered trademark) Human Genome U133 Plus 2.0 Array is an array that can detect the expression of more than 47,000 genes present on the genome. The method of use is to amplify a sample (total RNA, mRNA, etc.) by PCR using one T7-Oligo (dT) primer and label it. One array is designed to be complementary for each gene expression, and after hybridization, the gene expression of the sample is determined based on the signal, and the expression level of each gene is determined by the signal intensity or Comparisons can be made based on signal ratio. For details of this DNA chip, refer to the product information published at http://www.affymetrix.com/products_services/arrays/specific/hgu133plus.affx.

In addition to the above, the gene expression level can be measured by any method known to those skilled in the art. Examples of such a method include a method using a primer specific for gene expression and a method using quantitative PCR (qPCR).
In this method, it is preferable to use a DNA chip (microarray) capable of measuring gene expression simply and rapidly.

cDNA synthesis Synthesis of cDNA from nuclear RNA or mRNA can be performed according to a method known in the art (Japanese Patent No. 4111985). Here, “nuclear RNA” refers to a primary transcript that has not been spliced after being transcribed from genomic DNA, and therefore does not migrate to the cytoplasm and is still present in the nucleus. Say. That is, many nuclear RNAs contain both exons and introns on the genome and have a long chain length. Design of a primer for synthesis or amplification of a specific gene can be performed according to a general primer design method known in the art.

DNA extraction Genomic DNA can be extracted using a method known in the art (such as a phenol / chloroform method) or a commercially available kit.

Typing of genetic polymorphism in cDNA or genomic DNA
Typing (detection) of gene polymorphism (SNP or haplotype, etc.) in cDNA or genomic DNA can be performed using techniques known in the art. For example, gene polymorphism typing can be performed by hybridization with a probe specific to one gene polymorphism. The probe can be labeled by an appropriate means such as a fluorescent substance or a radioactive substance, if necessary. The probe may be any probe as long as it contains a gene polymorphic site and specifically hybridizes with cDNA, and specific probe designs are known in the art. The hybridization conditions may be any conditions sufficient to distinguish gene polymorphisms. For example, hybridization occurs in the case of one gene polymorphism, but hybridization occurs in the case of another gene polymorphism. Such as stringent conditions, such conditions are known to those skilled in the art.

  The probe can be used as a DNA chip (microarray) with one end fixed to a substrate. In this case, only a probe corresponding to one gene polymorphism may be immobilized on the DNA chip, or probes corresponding to both gene polymorphisms may be immobilized. The detection of genetic polymorphism using such a DNA chip is described in, for example, “DNA microarray and latest PCR method”, supervised by Masaaki Muramatsu and Hiroyuki Nami, Shujunsha, 2000, Chapter 10.

  As a specific example of gene polymorphism detection using a DNA chip, a method using a GeneChip (registered trademark) Human Mapping 500K array manufactured by Affymetrix will be described. The GeneChip (registered trademark) Human Mapping 500K array is an array that includes two arrays and can detect more than 500,000 SNPs present on the genome. The method of use is to cut the sample (genome, cDNA, etc.) with a restriction enzyme (Sty or Nsp), attach an adapter, and use one type of primer specific to that adapter (one for Sty or Nsp). Amplify by PCR reaction and label. The two arrays are designed to be complementary for each allele of each SNP.After hybridization, the SNP of the sample is judged based on the signal, and the expression level of each allele is converted to the signal intensity or signal ratio. (The detailed method is described in the patent application (PCT / JP2005 / 023439 or the above-mentioned Japanese Patent Number; 4111985, Method for identifying a gene with allellic variation in gene expression. FILING DATA: December 15, 2005 Year))). For details of this DNA chip, refer to the product information published at http://www.affymetrix.com/jp/products_services/arrays/specific/500k.affx.

  Moreover, gene polymorphism can be type | molded by all the methods well-known to those skilled in the art besides the above-mentioned. Such methods include using primers specific for gene polymorphism, using restriction fragment length polymorphism (RFLP), direct sequencing, denaturing gradient gel electrophoresis (DGGE), mismatch site A method using chemical cleavage (CCM), a primer extension method (TaqMan method), or the like can be used.

  In this method, it is preferable to use a DNA chip (microarray) that can type gene polymorphisms simply and quickly.

The present invention includes a method for searching gene polymorphisms involved in sorafenib stimulation responsiveness from data of ExpressGenotyping method, and the method has the following steps.
(i) (b) By comparing the result of ExpressGenotyping method in which only solvent is exposed to cells and the result of ExpressGenotyping method in which drug is exposed to cells (b), markers that are responsive to drug stimulation SNPs that show differences between alleles in the gene expression level to be selected.
(ii) Data processing is performed using a specific algorithm on a computer that compares the results of the EG method of (a) and (b).

  By the above method, genes whose expression levels differ between alleles with respect to the searched sorafenib stimulation, that is, genes that differ in expression among alleles specifically for sorafenib stimulation, and polymorphisms thereof, that is, fluctuations in expression between alleles due to sorafenib stimulation. Influencing polymorphisms can be used to determine the responsiveness to sorafenib in a subject receiving sorafenib. In addition, the polymorphism can be used to analyze the relationship between the responsiveness of the subject at the time of stimulation with sorafenib and the polymorphism.

  According to the above method, a gene polymorphism of a gene whose expression level differs between alleles with respect to the searched sorafenib stimulation can be selected as a gene polymorphism for determining a gene whose expression level differs between alleles. There is a high possibility that a gene polymorphism capable of determining a gene whose expression level differs between alleles is related to a phenotype than other gene polymorphisms. Only by detecting the gene polymorphism, it is possible to examine whether or not a certain specimen has an allele with a high expression level. Genes with different expression levels between alleles are associated with a phenotype of responsiveness to drugs. The frequency of having a particular genetic polymorphism is significantly higher in subjects exhibiting a particular phenotype (sensitivity to sorafenib). The difference in the gene polymorphism affects the quantitative regulation of the expression level of the gene involved in the phenotype.

  As a result of the above search, 3224 genes (6953 probe) were searched as genes having different expression levels between alleles and single nucleotide polymorphisms of each gene by sorafenib stimulation.

Among these 3224 genes, the following 10 genes in particular were selected as genes whose expression levels differ greatly between alleles in response to sorafenib stimulation.
(a) dihydropyrimidine dehydrogenase (DPYD) gene,
(b) CD247 molecule gene,
(c) phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene,
(d) interleukin 1 receptor, type II (IL1R2) gene,
(e) dedicator of cytokinesis 2 (DOCK2) gene,
(f) estrogen receptor 1 (ESR1) gene,
(g) complement component 5 (C5) gene,
(h) phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene,
(i) the pancreatic lipase-related protein 3 (PNLIPRP3) gene, and
(j) lipase, hepatic (LIPC) gene.

In the genes (a) to (j), 52 polymorphisms (SNPs) were selected as gene polymorphisms that can determine the difference in expression level between alleles of each gene.
5 to 14 show the positions of polymorphisms in the base sequences of the genes (a) to (j), respectively.

In the present invention, gene polymorphisms of the genes (a) to (j) to be typed include the following.
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLCB1) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, the single nucleotide polymorphism rs1431485, the gene polymorphism present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of
(x) single nucleotide polymorphisms in lipase, hepatic (LIPC) genes rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542, and gene polymorphisms in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in

  Here, the gene polymorphism present in the vicinity of the gene polymorphism refers to a gene polymorphism located within 30,000 kb, preferably within about 10,000 kb of the drug from the gene polymorphism. Such polymorphisms located in the vicinity are highly likely to be recombined together during chromosome recombination. The gene polymorphism in linkage disequilibrium with the gene polymorphism is a gene polymorphism related to the gene polymorphism. Specifically, when the gene polymorphism is X, It is such that the relationship that another genetic polymorphism is always Y is established. Analysis of the relationship between the responsiveness of a subject at the time of sorafenib stimulation using a polymorphism and the polymorphism is, for example, the above gene in a subject known to exhibit a specific responsiveness to the sorafenib stimulation in advance. You can check the polymorphism of. Moreover, the polymorphism of the said gene in the subject which is going to administer sorafenib is investigated beforehand, the responsiveness when sorafenib is administered can be investigated, and a polymorphism and responsiveness can also be linked | related. These can be performed in vivo, but a subject's cells can be collected and isolated, and the isolated cells can be used for association in vitro. Furthermore, the responsiveness to the sorafenib stimulation in the cultured cell which can be obtained can be investigated in vitro, and it can also relate by investigating the polymorphism of each cultured cell. For example, by examining the relationship between the polymorphism of the 16th base in the nucleotide sequence shown in SEQ ID NO: 1 of the dihydropyrimidine dehydrogenase gene and the response to sorafenib stimulation, side effects of sorafenib are observed in subjects whose polymorphic site is C. If a large phenotype appears, the polymorphic site is associated with the genotype C and a phenotype with significant side effects of sorafenib. Sorafenib administration can be discontinued.

  The present invention compares (b) the result of ExpressGenotyping method in which only cells are exposed to a solvent and (b) the result of ExpressGenotyping method in which a drug is exposed to cells, thereby providing a marker responsive to drug stimulation (depending on drug exposure). A method of associating responsiveness with SNPs that show differences between alleles in the gene expression level that is manifested is also included.

  In the present invention, at least one of the above 10 genes is used. A plurality of genes may be used at once, for example, 2, 3, 4, 5, 6, 7, 8, 9 or 10 of the above 10 types of genes may be used. In addition, gene polymorphism can be determined using at least one polymorphism of 52 polymorphisms of 10 types of genes, and a plurality of polymorphisms may be used at once. For example, among 52 polymorphisms, 2 or more, 3 or more, 4 or more, 5 or more, 10 or more, 15 or more, 20 or more, 25 or more, 30 or more, 35 or more, 40 or more, 45 or more, 50 or more, 51 or more Alternatively, determination can be made using 52.

  The determination using the gene and its polymorphism means that the polymorphism of the gene in a subject to be administered sorafenib is measured in advance, and the response to sorafenib is predicted by the type of base at the polymorphic site, or It means to evaluate and judge. Responsiveness to sorafenib refers to the type of side effects that can occur due to sorafenib, the degree of side effects that can occur, and the degree of efficacy of sorafenib. It has been known that there is an individual difference in these responsiveness, and conventionally, the individual difference could not be predicted and evaluated, but can be predicted and evaluated by the method of the present invention.

  That is, the polymorphism of the gene can be used as a biomarker for determining the responsiveness of a subject to sorafenib before administration of sorafenib.

  Polymorphism detection includes detection of one chromosome on a pair of chromosomes as well as detection of both chromosomes, and even when detecting both chromosomes, homozygosity at a single nucleotide polymorphism site. Or detection of heterozygosity.

  Furthermore, the present invention includes an oligonucleotide or a label thereof used for detecting the above polymorphism, and the oligonucleotide or the label thereof can be used as a probe or a primer. These oligonucleotides consist of a DNA fragment comprising a base sequence containing the single nucleotide polymorphic site of the above gene or a base sequence complementary to the base sequence, and such oligonucleotides are used for detecting single nucleotide polymorphisms. Can be used as a probe. In addition, a base sequence in the vicinity of or away from the gene polymorphic site can be used as a primer for amplifying the base sequence containing the gene polymorphic site. In this case, two types of sequences existing on the 3 ′ side and 5 ′ side of the polymorphic site can be used as primer pairs. The number of bases constituting the oligonucleotide used for detecting the polymorphism is 5 to 50, preferably 10 to 30, more preferably 15 to 25. From the continuous base sequence including the polymorphic site of the base sequence of the above gene, Become. In addition, oligonucleotides having several mismatches, preferably 1 to 5, more preferably 1 or 2, and particularly preferably 1 mismatch in the continuous base sequence including the polymorphic site of the base sequence of the above gene. Can be used. Polymorphism can be detected by a hybridization assay using a probe. The oligonucleotide of the present invention can be prepared by chemical synthesis or can be prepared as an amplification product obtained by amplifying a gene by PCR using the above primers. The probe of the present invention may be labeled with a fluorescent substance, an enzyme, a radioisotope, a chemiluminescent substance or the like for detection. As a labeling substance used for labeling, a known substance can be used and labeled by a known method. Examples of the fluorescent substance include Cy3, Cy5, rhodamine, fluorescein and the like.

  Furthermore, the present invention includes an immobilized substrate on which the oligonucleotide is immobilized. Various substrates such as a slide glass, a nitrocellulose membrane, and microbeads can be used as the substrate on which the oligonucleotide is immobilized. When a plurality of oligonucleotides are aligned and immobilized on an immobilized substrate, the immobilized substrate can be used as a DNA microarray or a DNA chip. The oligonucleotide may be synthesized on a substrate, or the synthesized oligonucleotide may be immobilized on the substrate. Immobilization on a substrate can be performed using, for example, a commercially available spotter or arrayer, and oligonucleotides can be immobilized by adsorption or binding via covalent bond, and by covalent bond binding. In the case of immobilization, functional groups such as an amino group for covalent bonding and an SH group may be introduced into the substrate surface and the oligonucleotide.

  Polymorphism detection using an immobilized substrate on which an oligonucleotide is immobilized can be performed by a known method.

  The present invention will be specifically described by the following examples, but the present invention is not limited to these examples.

Example 1 Search for Genes with Different Expression Levels Between Alleles In this example, genes with different expression levels between alleles were searched for by sorafenib stimulation.

  Sorafenib stimulation was performed using 10 lymphocytes (Table 1) established by EB virus. Ten specimens of lymphocytes were purchased from Coriell Institute for Medical Research (Coriell), passaged once, and then stored in a cell storage (-80 ° C storage). The group composition in the sorafenib stimulation was 1) the control group (DMSO), 2) the sorafenib stimulation group (2 groups of drug stimulation on / off), the concentration of sorafenib was 10 μM, and the DMSO concentration of the solvent was 0.01% (The optimal exposure conditions were determined from the results of mRNA expression analysis (FIG. 1)).

Start of sorafenib stimulation: 5% at 37 ° C in RPMI medium 1640 (Invitrogen) medium supplemented with 15% Fetal Bovine Serum (Invitrogen) until the number of cells reaches the number that can be used in the test (1.25 x 10 ⁷ cells or more) The cells were cultured under CO ₂ conditions. When the number of cells capable of stimulating sorafenib was reached, two culture dishes (Nalge Nunc International) for 10-20 ml were prepared and seeded so that each dish had 5 × 10 ⁶ cells. Next, DMSO was added to one dish where cells were seeded so that the DMSO concentration was 0.1%, and sorafenib was added to the other dish so that the stimulating concentration of sorafenib was 10 μM. Thereafter, the cells were cultured in RPMI medium 1640 medium (without Fetal Bovine Serum) at 37 ° C. and 5% CO ₂ for 4 hours. Table 2 shows the grouping of stimulation conditions for sorafenib stimulation.

  RNA extraction from the DMSO group and the sorafenib-stimulated group was performed with TRIzol Reagent (Invitrogen). The concentration of the extracted RNA was measured with NanoDrop ND-1000 (NanoDrop Technologies), and the purity was evaluated with an Agilent 2100 bioanalyzer (Agilent Technologies).

RNA extracted from the DMSO group and the sorafenib-stimulated group was used for comprehensive measurement of the expression level and total amount of all Premature RNA alleles.
DNA extraction from the DMSO group and the sorafenib-stimulated group was performed with the QIAamp DNA Mini Kit (QIAGEN). The concentration of the extracted DNA was measured with NanoDrop ND-1000 (NanoDrop Technologies). DNA extracted from this DMSO group and sorafenib-stimulated group was used for SNP typing of DNA.

  Using the RNA of DMSO group and sorafenib-stimulated group of 10 lymphocytes, genes having different expression levels between alleles were determined specifically by the sorafenib-stimulated group by the method described in WO2006 / 068111.

  As a result, a shape close to a normal distribution curve considered to be noise appeared at a low signal ratio, and there were many signals (probes) based on gene polymorphisms existing in a region without a gene. From this frequency distribution and the positional relationship between the probe and the gene on the genome, the signal and noise derived from the gene (cDNA) can be separated by this assay, and the signal ratio is high by measuring the signal ratio between the cDNA and the genome. It was found that the signal may be considered as a signal from the gene that expressed the part (mainly cDNA derived from nuclear RNA).

  Accurate signal of cDNA of Sorafenib stimulation group to search for gene polymorphisms that determine genes whose expression level differs among alleles specifically from Sorafenib stimulation group from cDNA signal intensity that affects the expression level of DMSO group and Sorafenib stimulation group Take the ratio of intensity / accurate signal intensity of cDNA in the DMSO group (EAI; Expression Allelic Imbalance, expression level between alleles is unbalanced / different), and probe with a ratio of 1.5 times or more sorafenib It was narrowed down as genes (gene polymorphisms) whose expression levels differed between alleles specifically for the stimulation group (details are described in [Example 2]).

  Thus, when the difference in the expression level between one allele and the other allele was compared using the EAI of the sorafenib-stimulated group, 3224 genes (6953 probe) were significantly significant in the sorafenib-stimulated group. Showed the difference. Among them, the following 10 genes were selected as gene polymorphisms having different expression levels among alleles related to the side effect of sorafenib stimulation (FIG. 4).

(1) DPYD: dihydropyrimidine dehydrogenase
The protein of the DPYD gene is a pyrimidine-metabolizing enzyme, and it has been reported that mutations in this gene increase the risk of side effects in cancer patients in 5-fluorouracil (5-FU) chemotherapy. The fatal drug interaction "soribudine toxic injury" that occurred between the 5-FU anticancer drug and the anti-herpetic drug sorivudine is well known (Watabe T. et al., Yakugaku Zasshi. 2002 Aug; 122 (8): 527-35. PMID: 12187768, 2002).

Single nucleotide polymorphisms of DPYD gene that showed EAI by sorafenib stimulation rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887, and rs10875097 sequence information (GeneChip (registered trademark) Human Mapping 500K array (Affymetrix)) It is.
rs10449721:
agcagcttcaatggaa [C / T] gatgagaagtagaatc (SEQ ID NO: 1)
rs10875047:
cgtgacaaggaagacc [A / G] agatggaatatatgag (SEQ ID NO: 2)
rs11165805:
ggaataatagtcattg [C / G] agtctcaaaacggtgg (SEQ ID NO: 3)
rs11165867:
ataatctgctcttaga [A / G] gactataaccccataa (SEQ ID NO: 4)
rs6687374:
ccacaccaccctaaac [A / G] agtcatcatagtagaa (SEQ ID NO: 5)
rs4949952:
ggagcccagttacgaa [A / G] ggccgtttatgcttgc (SEQ ID NO: 6)
rs11165887:
aatccatgagtgagaa [A / G] aactaatatcatgtga (SEQ ID NO: 7)
rs10875097:
ctatagtaagtgtcca [C / T] gttaaggtgtttgata (SEQ ID NO: 8)

(2) CD247: CD247 molecule
The protein of the CD247 gene is a T cell receptor (zeta) and plays an important role in recognizing antigens (Minguet S. et al., Immunol Lett. Mar 15; 116 (2): 195-202. Dec 20; 7 (14): 1909-23. PMID: 18207249, 2008).

The sequence information of the single nucleotide polymorphism rs864537 (GeneChip (registered trademark) Human Mapping 500K array (manufactured by Affymetrix)) of the CD247 gene that showed EAI upon stimulation with sorafenib is as follows.
rs864537:
agtgtttttatttgga [A / G] catcattgcactgtga (SEQ ID NO: 9)

(3) PLA2G4A: phospholipase A2, group IVA (cytosolic, calcium-dependent)
The protein of the PLA2G4A gene belongs to the group IV family of phospholipase A2 and is a catalyst for releasing arachidonic acid from membrane phospholipids (lipid-derived substances such as arachidonic acid regulate hemodynamics and inflammation (Rubin BB. Et al., J Biol Chem. Mar 4; 280 (9): 7519-29. Epub 2004 Oct 8. PMID: 15475363, 2005).

The sequence information (GeneChip (registered trademark) Human Mapping 500K array (Affymetrix)) of single nucleotide polymorphisms rs10752979, rs10798069, and rs10911963 of the PLA2G4A gene that showed EAI by sorafenib stimulation is as follows.
rs10752979:
tgggcctgaaatacaa [C / T] gagaaaatgctcttga (SEQ ID NO: 10)
rs10798069:
catcctgccagagaga [A / C] aatttttgtgtaaagg (SEQ ID NO: 11)
rs10911963:
agaagcttaacaatga [A / G] ggttaatgatgcacca (SEQ ID NO: 12)

(4) IL1R2: interleukin 1 receptor, type II
The IL1R2 gene protein is well known as a cytokine receptor, and binds to interleukin alpha (IL1A), interleukin beta (IL1B), and interleukin 1 receptor type I (IL1R1 / IL1RA) as a decoy that inhibits the activity of the ligand. Working (Re F. et al., J Exp Med. Feb 1; 179 (2): 739-43. PMID: 8294881, 1994).

The sequence information of the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920 (GeneChip (registered trademark) Human Mapping 500K array (manufactured by Affymetrix)) of the IL1R2 gene that showed EAI by stimulation with sorafenib is as follows.
rs4851527:
acagctagtaagcaga [C / T] gttgaaatttaaaccc (SEQ ID NO: 13)
rs3218872:
tcctactccttcttga [C / T] ggatgaagttttaggc (SEQ ID NO: 14)
rs3218920:
cggtctacataagcaa [C / T] atctaatgacaatatc (SEQ ID NO: 15)

(5) DOCK2: dedicator of cytokinesis 2
DOCK2 gene protein is a CDM family of hematopoietic cells essential for lymphocyte chemotaxis (Reif K. et al., Trends Cell Biol. Aug; 12 (8): 368-73. PMID: 12191913, 2002) Year).

Single nucleotide polymorphisms of DOCK2 gene that showed EAI by sorafenib stimulation rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638 sequence information (GeneChip (registered trademark) Human Mapping 500K array (Affymetrix)) It is.
rs900464:
gttttaaagatggaaa [C / G] gatgtggatatgtaga (SEQ ID NO: 16)
rs264846:
aggcctggattaagaa [A / T] gaaatcaaacaaggta (SEQ ID NO: 17)
rs90213:
ctaaatagtgttagga [C / T] gttcagttattcagtt (SEQ ID NO: 18)
rs966595:
gccataaaattgacta [C / T] agccagtgtccttcaa (SEQ ID NO: 19)
rs4867895:
ctgctgcctaccaaat [G / T] aagtcataaatgcatt (SEQ ID NO: 20)
rs11134600:
atgaggtaccaatacc [A / G] catactgggaaatata (SEQ ID NO: 21)
rs2287727:
ggatggagaagccaat [G / T] agccgtctcatgtccc (SEQ ID NO: 22)
rs1316638
ctttcttcttcttacc [A / G] tcctatgtacagcttc (SEQ ID NO: 23)

  Furthermore, the phenotype of this DOCK2 gene polymorphism (rs11134600) was examined. In general, the side effects of sorafenib differed between races (increased in the order of Japanese> Caucasian> black), and we examined whether there are differences between races due to this DOCK2 gene polymorphism. Allele frequency data between races (specific data of Entrez SNP: http://www.ncbi.nlm.nih.gov/sites/entrez?db=snp&cmd=search&term) (See =). As a result, it was found that the above-mentioned DOCK2 rs11134600 is a genetic polymorphism having different degrees of influence by sorafenib stimulation (FIG. 3).

  From this result, it was considered that a side effect gene caused by sorafenib stimulation could be clarified as a future prospect by typing SNPs of DNA polymorphisms found in the present invention with DNA of clinical specimens.

(6) ESR1: estrogen receptor 1
The ESR1 protein is an estrogen receptor, a ligand-activated transcription factor that plays an important role in hormone and DNA binding and transcriptional activity. This estrogen receptor has also been implicated in the development of breast cancer, endometrial cancer and osteoporosis (Ali S. et al., J Mammary Gland Biol Neoplasia. Jul; 5 (3): 271-81. PMID: 14973389 , 2000; Wedren S. et al., BMC Cancer. Nov 6; 8: 322.PMID: 18990228, 2008; Ferrari S. et al., Best Pract Res Clin Endocrinol Metab. Oct; 22 (5): 723 -35. PMID: 19028354, 2008).

The sequence information of the single nucleotide polymorphisms rs9371557 and rs3798573 of the ESR1 gene that showed EAI by sorafenib stimulation (GeneChip (registered trademark) Human Mapping 500K array (manufactured by Affymetrix)) is as follows.
rs9371557:
gggtacatggtacaga [C / T] gaagatcactatgaag (SEQ ID NO: 24)
rs3798573:
tgccaaatccaaatga [C / T] ggacgttgcagaacac (SEQ ID NO: 25)

(7) C5: complement component 5
The protein of the C5 gene is a component of complement and plays an important role in the process of inflammation and cell death (DiScipio RG. Et al., Int Immunopharmacol. Dec 20; 7 (14): 1909-23. PMID: 18039528, 2007).

The sequence information of the single nucleotide polymorphism rs10985112 (GeneChip (registered trademark) Human Mapping 500K array (manufactured by Affymetrix)) of the C5 gene that showed EAI by sorafenib stimulation is as follows.
rs10985112:
ggatgctggcacataa [A / G] acaagaaaacaaacag (SEQ ID NO: 26)

(8) PLCB1: phospholipase C, beta 1 (phosphoinositide-specific)
The protein of the PLCB1 gene functions as a catalyst in the formation of inositol 1,4,5-triphosphate and diacylglycerol from phosphatidylinositol-4,5-diphosphate. It plays an important role in signal transduction of extracellular stimuli into cells (Fantuzzi L. et al., Blood. Apr 1; 111 (7): 3355-63. PMID: 18203956, 2008).

Single nucleotide polymorphisms of PLCB1 gene that showed EAI by sorafenib stimulation rs2235212, rs6055624, rs2294257, rs2180532, rs10485723, rs1156958, rs1237829, rs2050090, rs6055925, rs6039215, rs6140677, rs6133610, rs6133621, rs13040543, rs10485727, 605 The sequence information of rs3848831 and rs6086678 (GeneChip (registered trademark) Human Mapping 500K array (manufactured by Affymetrix)) is as follows.
rs2235212:
ttaccagaaattaaag [C / T] caccagagtggttgct (SEQ ID NO: 27)
rs6055624:
cttgattagcttacta [C / T] gtactggctgactaca (SEQ ID NO: 28)
rs2294257:
cccttctagccaaaaa [C / T] gtgttcagttagacat (SEQ ID NO: 29)
rs2180532:
aaaaaagccatgaaag [A / T] ctgatatttcctagaa (SEQ ID NO: 30)
rs10485723:
catagaaggttaccta [C / T] atcattgccttgacct (SEQ ID NO: 31)
rs1156958:
tattttaccggtaccc [C / T] atagtcctcaaaatca (SEQ ID NO: 32)
rs1237829:
tctggtgtaatttcaa [C / T] gtaaatccaatttgtt (SEQ ID NO: 33)
rs2050090:
caggtcaggttttgac [A / G] gctaatgaattataga (SEQ ID NO: 34)
rs6055925:
gaggggtattaatgaa [C / G] tgtcaaatatacatag (SEQ ID NO: 35)
rs6039215
gaaagcagttcgacaa [A / T] cttaaaactgcattac (SEQ ID NO: 36)
rs6140677:
taaagaaatgcgttca [A / G] taagcatcaggtactt (SEQ ID NO: 37)
rs6133610:
gcttgaaaaccactgc [A / G] attgattaaggtaagt (SEQ ID NO: 38)
rs6133621:
catctagacagcttga [C / T] aaatggcatgactaca (SEQ ID NO: 39)
rs13040543:
tgtactgctaagcctc [A / G] aaaatccaagctgaaa (SEQ ID NO: 40)
rs10485727:
taaaggctattgccaa [C / T] gtttccagagctttca (SEQ ID NO: 41)
rs6118343:
ctgtaaaaaactaata [C / T] gctaacaacgaattct (SEQ ID NO: 42)
rs6056198:
caggacaggctagtaa [A / C] acaagatatgcctgga (SEQ ID NO: 43)
rs6086627:
atggagagtaacatgg [A / G] aagtttaaccaggtgg (SEQ ID NO: 44)
rs3848831:
atttggatctacttag [C / T] attttgcctacctcgt (SEQ ID NO: 45)
rs6086678:
tacccagtcccaataa [A / T] ttaaagcaaaaactgt (SEQ ID NO: 46)

(9) PNLIPRP3: pancreatic lipase-related protein 3
PNLIPRP3 is highly expressed in hepatocellular carcinoma (Saelee P. et al., Asian Pac J Cancer Prev. Jul-Sep; 10 (3): 501-6. PMID: 19640199, 2009).

The sequence information of the single nucleotide polymorphism rs1431485 (GeneChip (registered trademark) Human Mapping 500K array (manufactured by Affymetrix)) of the PNLIPRP3 gene that showed EAI by stimulation with sorafenib is as follows.
rs1431485:
tttaagagcctttgca [C / T] ggttccacagttgcat (SEQ ID NO: 47)

(10) LIPC: lipase, hepatic
LIPC is a hepatic triglyceride lipase and is expressed in the liver. LIPC functions as a coordinating and crosslinking factor when hydrolyzing triglycerides and taking up lipoproteins by receptors. LIPC has also been reported as a risk factor for type 2 diabetes (Todorova B. et al., J Clin Endocrinol Metab. May; 89 (5): 2019-23. PMID: 15126514, 2004).

The sequence information of the single nucleotide polymorphisms rs12438071, rs4775047, rs9920144, rs7179747, and rs3751542 of the LIPC gene that showed EAI by sorafenib stimulation (GeneChip (registered trademark) Human Mapping 500K array (Affymetrix)) is as follows.
rs12438071:
tcttgatactactaag [C / T] caccgaggagccgtgg (SEQ ID NO: 48)
rs4775047:
aatgattgagtttcaa [C / T] gtgtccttatttcatt (SEQ ID NO: 49)
rs9920144:
gtggcatatcgtaaac [A / G] gggtgccttgtaggcc (SEQ ID NO: 50)
rs7179747:
gaacttgggcctaaaa [G / T] ctttctgttttatcac (SEQ ID NO: 51)
rs3751542:
gagtattattaggccc [C / T] agtgatggagtacaga (SEQ ID NO: 52)

Example 2 Method for Searching for Gene Polymorphisms Involved in Sorafenib Stimulus Responsiveness In this example, a method for searching for gene polymorphisms involved in sorafenib stimulus responsiveness was developed from the data of ExpressGenotyping method.

In order to select SNPs that show differences between alleles of gene expression levels that are manifested by drug exposure, the exact signal intensity of the cDNA of the sorafenib-stimulated group [(b) Results of the EG method in which the drug was exposed to cells] / DMSO group Calculate the ratio of the exact signal intensity of the cDNA [(b) Result of EG method in which cells were exposed to solvent only], and the expression level of the probe with a ratio of 1.5 times or more differs between alleles specifically for the sorafenib stimulation group Narrowed down as genes (gene polymorphisms showing EAI). The data processing method is described below.
(i) The results of (b) 10 samples of DMSO_nsp_sty file and (b) 10 samples of Sorafenib_nsp_sty file processed by the EG algorithm were obtained as txt files.
(ii) The files (a) and (b) were combined on Microsoft Office Access 2007 (Microsoft) using Probe ID.
(iii) Only probes with a heterozygous call (AB call) were obtained from each of the combined files (using Microsoft Office Excel 2007 (Microsoft)).
(iv) Only the gene regions (3UTR, 5UTR, CDS, exon, and intron) were obtained from each linked file (using Microsoft Office Excel 2007 (Microsoft)).
(v) Calculate the ratio of (b) Sora AI [adj ratio (upper) / (b) adj ratio (lower)] / DMSO AI [adj ratio (upper) / adj ratio (lower)] with each file combined Then, probes with a ratio of 1.5 or more were obtained as specific EAI probes in the sorafenib stimulation group (using Microsoft Office Excel 2007 (Microsoft)).
(vi) The EAI probe files for 10 samples were combined on Microsoft Office Access 2007 (Microsoft), and the number of EAI probes was counted (probe ID was used for binding).

  By the above method, genes (gene polymorphisms) having different expression levels between alleles were obtained specifically for the sorafenib stimulation group.

  Using the method, probe, and substrate of the present invention, it is possible to predict different responsiveness for each individual of sorafenib.

SEQ ID NOs: 1 to 52

Claims (15)

Differences in expression levels between alleles caused by sorafenib stimulation In vitro detection of gene polymorphisms that can determine differences in expression levels in any of the genes (a) to (j) below. Methods for predicting responsiveness of sorafenib in an administered subject:
(a) the dihydropyrimidine dehydrogenase (DPYD) gene;
(b) the CD247 molecule gene;
(c) phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene;
(d) interleukin 1 receptor, type II (IL1R2) gene;
(e) dedicator of cytokinesis 2 (DOCK2) gene;
(f) estrogen receptor 1 (ESR1) gene;
(g) complement component 5 (C5) gene;
(h) phospholipase C, beta 1 (phosphoinositide-specific) (PLBC1) gene;
(i) the pancreatic lipase-related protein 3 (PNLIPRP3) gene; and
(j) lipase, hepatic (LIPC) gene. The method for predicting responsiveness of sorafenib in a subject to which sorafenib is administered according to claim 1, wherein the gene polymorphism capable of determining the difference in expression level in (a) to (j) is any of the following:
(I) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any one of SEQ ID NOs: 1 to 8 in the dihydropyrimidine dehydrogenase (DPYD) gene;
(ii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 9 in the CD247 molecule gene;
(iii) A single nucleotide polymorphism present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 10 to 12 in the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene Site;
(iv) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 13 to 15 in the interleukin 1 receptor, type II (IL1R2) gene;
(v) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 16 to 23 in the dedicator of cytokinesis 2 (DOCK2) gene;
(vi) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 24 or 25 in the estrogen receptor 1 (ESR1) gene;
(vii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 26 in the complement component 5 (C5) gene;
(viii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 27 to 46 in the phospholipase C, beta 1 (phosphoinositide-specific) (PLBC) gene;
(ix) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 47 in the pancreatic lipase-related protein 3 (PNLIPRP3) gene; or
(x) A single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 48 to 52 in the lipase, hepatic (LIPC) gene. The method for predicting responsiveness of sorafenib in a subject to which sorafenib is administered according to claim 1, wherein the gene polymorphism capable of determining the difference in expression level in (a) to (j) is any of the following:
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLBC) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, a single nucleotide polymorphism rs1431485, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of types, or
(x) single nucleotide polymorphisms in lipase, hepatic (LIPC) genes rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542, and gene polymorphisms in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in   The response of sorafenib in a subject to which sorafenib is administered according to any one of claims 1 to 3, wherein the prediction of the response of sorafenib is a prediction of the effect of sorafenib or a prediction of the type or degree of side effects in the subject. How to predict gender. Any of the following oligonucleotides comprising a DNA fragment containing a single nucleotide polymorphic site of (i) to (x), the partial sequence consisting of 10 to 30 bases of the following genes (i) to (x) Alternatively, a probe for predicting the response of sorafenib in a subject to which sorafenib is administered, comprising an oligonucleotide having a sequence complementary to the partial sequence or a label thereof:
(I) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any one of SEQ ID NOs: 1 to 8 in the dihydropyrimidine dehydrogenase (DPYD) gene;
(ii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 9 in the CD247 molecule gene;
(iii) A single nucleotide polymorphism present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 10 to 12 in the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene Site;
(iv) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 13 to 15 in the interleukin 1 receptor, type II (IL1R2) gene;
(v) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 16 to 23 in the dedicator of cytokinesis 2 (DOCK2) gene;
(vi) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 24 or 25 in the estrogen receptor 1 (ESR1) gene;
(vii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 26 in the complement component 5 (C5) gene;
(viii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 27 to 46 in the phospholipase C, beta 1 (phosphoinositide-specific) (PLBC) gene;
(ix) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 47 in the pancreatic lipase-related protein 3 (PNLIPRP3) gene; or
(x) A single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 48 to 52 in the lipase, hepatic (LIPC) gene. Any of the following oligonucleotides comprising a DNA fragment containing a single nucleotide polymorphic site of (i) to (x), the partial sequence consisting of 10 to 30 bases of the following genes (i) to (x) Alternatively, a probe for predicting the response of sorafenib in a subject to which sorafenib is administered, comprising an oligonucleotide having a sequence complementary to the partial sequence or a label thereof:
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and gene polymorphisms present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLBC) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, a single nucleotide polymorphism rs1431485, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of types, or
(x) single nucleotide polymorphisms rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542 in the lipase, hepatic (LIPC) gene, and gene polymorphisms present in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in   7. To predict sorafenib responsiveness in a subject to which sorafenib is administered according to claim 5 or 6, wherein the prediction of sorafenib responsiveness is prediction of the effect of sorafenib in a subject or prediction of the type or degree of side effects. Probe. An oligonucleotide consisting of a DNA fragment containing the following single nucleotide polymorphic sites (i) to (x): a partial sequence consisting of 10 to 30 bases of the following genes (i) to (x) or a portion thereof: An immobilized substrate for predicting responsiveness of sorafenib in a subject to which sorafenib is administered, to which at least one oligonucleotide having a sequence complementary to the sequence or a label thereof is immobilized:
(i) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 1 to 8 in the dihydropyrimidine dehydrogenase (DPYD) gene;
(ii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 9 in the CD247 molecule gene;
(iii) A single nucleotide polymorphism present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 10 to 12 in the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene Site;
(iv) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 13 to 15 in the interleukin 1 receptor, type II (IL1R2) gene;
(v) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 16 to 23 in the dedicator of cytokinesis 2 (DOCK2) gene;
(vi) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 24 or 25 in the estrogen receptor 1 (ESR1) gene;
(vii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 26 in the complement component 5 (C5) gene;
(viii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 27 to 46 in the phospholipase C, beta 1 (phosphoinositide-specific) (PLBC) gene;
(ix) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 47 in the pancreatic lipase-related protein 3 (PNLIPRP3) gene; or
(x) A single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 48 to 52 in the lipase, hepatic (LIPC) gene. An oligonucleotide consisting of a DNA fragment containing the following single nucleotide polymorphic sites (i) to (x): a partial sequence consisting of 10 to 30 bases of the following genes (i) to (x) or a portion thereof: An immobilized substrate for predicting responsiveness of sorafenib in a subject to which sorafenib is administered, to which at least one oligonucleotide having a sequence complementary to the sequence or a label thereof is immobilized:
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLBC) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, a single nucleotide polymorphism rs1431485, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of types, or
(x) single nucleotide polymorphisms in lipase, hepatic (LIPC) genes rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542, and gene polymorphisms in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in   10. To predict sorafenib responsiveness in a subject administered sorafenib according to claim 8 or 9, wherein the prediction of sorafenib responsiveness is a prediction of the effect of sorafenib in a subject or a prediction of the type or extent of side effects. Immobilization board. The following (i) to (x) are at least a pair of primer sets used for amplification of a DNA fragment containing a single nucleotide polymorphic site, and among the DNA polymorphic sites of the following genes (i) to (x) Sorafenib of a subject to which sorafenib is administered, comprising an oligonucleotide consisting of a partial sequence consisting of 10 to 30 bases present on the 3 ′ side and 5 ′ side of at least one polymorphic site or a sequence complementary to the partial sequence A pair of primer sets to predict the responsiveness of:
(i) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 1 to 8 in the dihydropyrimidine dehydrogenase (DPYD) gene;
(ii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 9 in the CD247 molecule gene;
(iii) A single nucleotide polymorphism present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 10 to 12 in the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene Site;
(iv) a single nucleotide polymorphic site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 13 to 15 in the interleukin 1 receptor, type II (IL1R2) gene;
(v) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by any of SEQ ID NOs: 16 to 23 in the dedicator of cytokinesis 2 (DOCK2) gene;
(vi) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 24 or 25 in the estrogen receptor 1 (ESR1) gene;
(vii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 26 in the complement component 5 (C5) gene;
(viii) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 27 to 46 in the phospholipase C, beta 1 (phosphoinositide-specific) (PLBC) gene;
(ix) a single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NO: 47 in the pancreatic lipase-related protein 3 (PNLIPRP3) gene; or
(x) A single nucleotide polymorphism site present at the position of the 16th base in the partial sequence represented by SEQ ID NOs: 48 to 52 in the lipase, hepatic (LIPC) gene. The following (i) to (x) are at least a pair of primer sets used for amplification of a DNA fragment containing a single nucleotide polymorphic site, and among the DNA polymorphic sites of the following genes (i) to (x) Sorafenib of a subject to which sorafenib is administered, comprising an oligonucleotide consisting of a partial sequence consisting of 10 to 30 bases present on the 3 ′ side and 5 ′ side of at least one polymorphic site or a sequence complementary to the partial sequence A pair of primer sets to predict the responsiveness of:
(i) single nucleotide polymorphisms in the dihydropyrimidine dehydrogenase (DPYD) gene rs10449721, rs10875047, rs11165805, rs11165867, rs6687374, rs4949952, rs11165887 and rs10875097, and the gene polymorphisms in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(ii) selected from the group consisting of a single nucleotide polymorphism rs864537, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism in the CD247 molecule gene At least one polymorph,
(iii) In the phospholipase A2, group IVA (cytosolic, calcium-dependent) (PLA2G4A) gene, the single nucleotide polymorphisms rs10752979, rs10798069 and rs10911963, the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the type,
(iv) In the interleukin 1 receptor, type II (IL1R2) gene, the single nucleotide polymorphisms rs4851527, rs3218872 and rs3218920, the gene polymorphisms present in the vicinity of the single nucleotide polymorphisms and linkage disequilibrium with the single nucleotide polymorphisms At least one polymorphism selected from the group consisting of a genetic polymorphism,
(v) single nucleotide polymorphisms in the dedicator of cytokinesis 2 (DOCK2) gene rs900464, rs264846, rs90213, rs966595, rs4867895, rs11134600, rs2287727 and rs1316638, and the gene polymorphism present in the vicinity of the single nucleotide polymorphism and the single nucleotide At least one polymorphism selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the base polymorphism,
(vi) From the single nucleotide polymorphisms rs9371557 and rs3798573 in the estrogen receptor 1 (ESR1) gene, the gene polymorphisms present in the vicinity of the single nucleotide polymorphism, and the gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of:
(vii) In the complement component 5 (C5) gene, a group consisting of a single nucleotide polymorphism rs10985112, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorph selected from,
(viii) Single nucleotide polymorphisms in phospholipase C, beta 1 (phosphoinositide-specific) (PLBC) gene Rs10485727, rs6118343, rs6056198, rs6086627, rs3848831 and rs6086678, and a genetic polymorphism present in the vicinity of the single nucleotide polymorphism and at least selected from the group consisting of genetic polymorphisms in linkage disequilibrium with the single nucleotide polymorphism One polymorph,
(ix) In the pancreatic lipase-related protein 3 (PNLIPRP3) gene, a single nucleotide polymorphism rs1431485, a gene polymorphism present in the vicinity of the single nucleotide polymorphism, and a gene polymorphism in linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of types, or
(x) single nucleotide polymorphisms in lipase, hepatic (LIPC) genes rs12438071, rs4775047, rs9920144, rs7179747 and rs3751542, and gene polymorphisms in the vicinity of the single nucleotide polymorphism and linkage disequilibrium with the single nucleotide polymorphism At least one polymorphism selected from the group consisting of genetic polymorphisms in   13. To predict sorafenib responsiveness in a subject to which sorafenib is administered according to claim 11 or 12, wherein the prediction of sorafenib responsiveness is prediction of the effect of sorafenib in a subject or prediction of the type or degree of side effects. A pair of primer sets.   The probe according to any one of claims 5 to 7, the immobilized substrate according to any one of claims 8 to 10, or the pair of primer sets according to any one of claims 11 to 13. A kit for predicting responsiveness of sorafenib in a subject to which sorafenib is administered.   The kit for determining the responsiveness of sorafenib in a subject to be administered sorafenib according to claim 14, wherein the prediction of responsiveness of sorafenib is a prediction of the effect of sorafenib or a prediction of the type or degree of side effects in the subject.

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