JP2011041553A - Marker for predicting effect of interferon therapy - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To identify an I-type interferon (IFN) response-associated gene for predicting therapeutic effect of a patient in IFN therapy, and provide a method for predicting the therapeutic effect of the I-type IFN therapy by analyzing the gene of the patient. <P>SOLUTION: A reagent for predicting sensitivity to the therapy with I-type IFN, containing a polynucleotide able to detect specific gene polymorphism presenting in a gene zone of human interferon λ2 (IFN-λ2) or interferon λ3 (IFN-λ3) and gene polymorphism having relation of linkage disequilibrium with the polymorphism is provided. The method for predicting sensitivity to the I-type IFN therapy by using the polymorphism of the gene of the patient as a marker is also provided. <P>COPYRIGHT: (C)2011,JPO&INPIT

Description

  The present invention relates to a marker for predicting a therapeutic effect of a patient in type I interferon therapy, its use, and the like.

Chronic hepatitis C is a viral disease caused by infection with hepatitis C virus (HCV), and interferon (IFN) therapy, which has the highest therapeutic effect, is used for its treatment. IFN therapy uses type I IFN, ie, IFN-α and IFN-β, specifically natural IFN-α and PEGylated IFN-α (peg IFN), etc. Is said to be high.
However, there are many patients who do not have a sufficient therapeutic effect with IFN-α monotherapy or combination therapy with IFN-α and antiviral agents (e.g. ribavirin), about 50% of patients infected with HCV genotype 1b, It has been reported that HCV is not excluded in about 20% of patients infected with HCV other than genotype 1b (see Non-Patent Documents 1, 2 and 3).

  Research into the causes of different treatment effects of IFN therapy for patients with chronic hepatitis C has been conducted energetically around the world, and viral and host (human) factors have been reported as factors affecting the therapeutic effect. . As a viral factor, it has been reported that when the HCV genotype 1b and the amount of virus in the blood before treatment are high, they are resistant to treatment (see Non-patent Documents 3 and 4).

  On the other hand, age, sex, race, and the like have been reported as host-side factors that affect the IFN treatment effect (see Non-Patent Document 5). In addition, it has been reported in individual gene regions that genetic polymorphisms such as cytokines are associated with the therapeutic effect of IFN therapy (see Non-Patent Documents 5 and 6).

  On the other hand, interferons λ2 (IFNλ2) and λ3 (IFNλ3) were identified as cytokines with unknown physiological activity and were named IL28A and IL28B, respectively. Later, these were new type III interferons, and it was found that there were three subtypes: λ1, λ2, and λ3. In addition, interferon λ has been reported to have an antiviral effect (see Non-Patent Documents 7 and 8). In addition, IFNλ2 and λ3 are known to bind to IL28Ra and IL10Rβ heteroreceptors (see Non-Patent Document 9). Furthermore, it has been reported that the expression of IFNλ2 or IFNλ3 is induced by a virus or IFN-α (see Non-Patent Document 10).

N. Engl. J. Med. 2002; 347: 975-982 NIH Consens Statement 1997; 15: 1-41 Ann. Intern. Med. 2000; 132: 296-305 J. Viral. Hepat. 2000; 7: 250-257 Hepatology 2004; 39: 880-890 J. Hepatol. 2002; 36: 271-277 Nature immunology 2003; 4 1: 69-77 J. Viral 2005; 79 6: 3851-3854 Nature immunology 2003; 4 1: 63-68 J. Viral 2006; 80 9: 4501-4509

  The problem to be solved by the present invention provides a method for predicting the therapeutic effect of IFN therapy by identifying an IFN-responsive gene for predicting the therapeutic effect of a patient in IFN therapy and analyzing the gene of the patient It is to be.

  In order to find a gene associated with the therapeutic effect of IFN therapy, the present inventors have conducted extensive studies focusing on genetic polymorphism, particularly single nucleotide polymorphism (SNP). SNP is defined as a point mutation with a frequency of 1% or more in a randomly selected population, and it exists as a good marker of genetic polymorphism because there is one on the gene every 1000 base pairs. Since recombination between homologous chromosomes does not occur between adjacent SNPs (linkage disequilibrium), the genetic diversity of multiple SNPs that cause linkage disequilibrium can be represented by a single selected SNP. SNP "(Nat. Rev. Cancer 2004, 4, 850-860; Nat. Genet. 2001, 29, 233-237). To illustrate, if SNP1 is an A or T allele and the adjacent SNP2 is a C or G allele, the SNP1 A allele and the SNP2 C allele are on the same chromosome. The case where there is a T allele of SNP1 and a G allele of SNP2 on the other homologous chromosome is set. If recombination does not occur between SNP1 and SNP2 during meiosis, the next generation will have the SNP1 A allele and SNP2 C allele on the same chromosome, and the SNP1 T allele and SNP2 G allele on the same chromosome. There are genes and there is an association between alleles (this is called linkage disequilibrium). In this case, if SNP1 is an A allele, SNP2 is a C allele, so these two SNPs can be represented by SNP1. In many cases, it has been demonstrated that there is a strong association between multiple adjacent SNPs, resulting in linkage disequilibrium, and polymorphisms in regions that cause linkage disequilibrium can be represented by tag SNPs. .

  In recent years, SNP analysis techniques and theories have developed dramatically, with a group of cases (groups suffering from diseases or groups with drug responsiveness) and a group of controls (groups of healthy individuals (or other patients suffering from other diseases) or drug responsiveness. By comparing the frequency of the tag SNP alleles between the two groups, the genes that are significantly different between the two groups are searched for, and genes related to disease and drug responsiveness (disease susceptibility) A case-control correlation analysis has been performed to identify genes or drug responsiveness related genes).

Therefore, the inventors of the present invention are Japanese patients suffering from chronic hepatitis C who are treated with IFN-α (HCV-infected patients), and more specifically, patients who are receiving peg IFN and rivapirin combination therapy. Genome-wide SNP genotyping was performed to extract two SNPs that correlate with the presence or absence of IFN therapy (P <10 ^-7 ). Since these SNPs were all located upstream of the IFNλ3 gene and showed strong linkage disequilibrium (r ² = 0.96), the SNP that gave the smallest P value was selected as the tag SNP.

For selected tag SNPs, (1) patients receiving IFN-α monotherapy, (2) patients receiving peg IFN and ribavirin combination therapy, and (3) IFN-α and ribavirin combination therapy. As a result of a case-control (IFN therapy effective group vs. non-response group) correlation analysis for each of the patients receiving this tag SNP, chronic C-type regardless of the mode of IFN therapy or HCV genotype Correlation with the therapeutic effect of IFN therapy for hepatitis was confirmed. Furthermore, the present inventors have identified about 42 kb of the vicinity of the SNP (including the gene regions of IFNλ2 and IFNλ3) for a considerable number of individuals in order to identify other genetic polymorphisms in linkage disequilibrium with the SNP. ) Was determined, and gene polymorphisms existing in the region were detected. As a result, 26 gene polymorphisms showing strong linkage disequilibrium with the tag SNP were identified. By testing the genotype of any of the identified gene polymorphisms, the patient's IFN treatment effect can be predicted. Further, although the identified SNP is likely to exist in the coding region of a gene that affects the IFN treatment effect of type I or in the vicinity thereof, the present inventors have identified the IFNλ2 and IFNλ3 genes as such genes.
The present invention has been completed based on the above findings.

That is, the present invention
[1] Gene polymorphisms existing in the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) gene regions located in human chromosome 19 short arm 19q13.13, and the following groups:
A) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 1;
B) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 2 (T>A);
C) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 3 (A>T);
D) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 4 (A>G);
E) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 5 (G>A);
F) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 6 (T>C);
G) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 7 (T>C);
H) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 8 (C>T);
I) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 9 (C>A);
J) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 10 (A>G);
K) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 11 (C>G);
L) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 12;
M) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 13 (A>G);
N) a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 14 (C>T);
O) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 15 (A>G);
P) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 16 (C>G);
Q) Gene polymorphism at the 301st base in the nucleotide sequence represented by SEQ ID NO: 17 (TT>G);
R) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 18 (G>C);
S) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 19 (C>A);
T) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 20 (C>T);
U) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 21 (->CT);
V) a gene polymorphism (C> T) at the 301st base in the nucleotide sequence represented by SEQ ID NO: 22;
W) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 23 (A>T);
X) a gene polymorphism (G> A) at the 301st base in the base sequence represented by SEQ ID NO: 24;
Y) gene polymorphism (-> GA) at the 301st base in the base sequence represented by SEQ ID NO: 25; and
Z) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 26 (AC>-);
(However, the parentheses indicate major allele> minor allele, and “-” indicates a base deletion.)
A reagent for predicting susceptibility to treatment with type I interferon, comprising a polynucleotide capable of detecting each minor allele in at least one gene polymorphism selected from
[2] The reagent according to [1], further comprising a polynucleotide capable of detecting each major allele,
[3] A polynucleotide capable of detecting each allele is a gene fragment of IFNλ2 or IFNλ3 consisting of 10 to 200 consecutive base sequences including the allele in the base sequence represented by each SEQ ID NO. The reagent according to [1] or [2] above, which is a probe capable of hybridizing under stringent conditions and / or a primer capable of amplifying the gene fragment,
[4] The reagent according to [3] above, wherein the gene fragment of IFNλ2 or IFNλ3 has a continuous base sequence of 10 to 50,
[5] The reagent according to [3] above, wherein the gene fragment of IFNλ2 or IFNλ3 comprises a continuous nucleotide sequence of 15 to 30,
[6] The reagent according to any one of [1] to [5], which is used for predicting whether or not interferon therapy is effective for patients with diseases to which treatment with type I interferon can be applied. ,
[7] The reagent according to [6] above, wherein the patient with a disease to which treatment with type I interferon can be applied is an HCV-infected patient,
[8] The reagent according to [7] above, wherein the HCV infected patient is an HCV genotype 1b infected patient, an HCV genotype 1a infected patient or an HCV genotype 2a infected patient,
[9] The reagent according to any one of [1] to [8] above, wherein the HCV-infected patient is East Asian.
[10] (1) A gene polymorphism present in the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) gene regions located in human chromosome 19 short arm 19q13.13, using a gene sample derived from a subject. The following groups:
A) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 1;
B) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 2 (T>A);
C) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 3 (A>T);
D) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 4 (A>G);
E) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 5 (G>A);
F) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 6 (T>C);
G) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 7 (T>C);
H) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 8 (C>T);
I) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 9 (C>A);
J) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 10 (A>G);
K) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 11 (C>G);
L) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 12;
M) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 13 (A>G);
N) a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 14 (C>T);
O) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 15 (A>G);
P) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 16 (C>G);
Q) Gene polymorphism at the 301st base in the nucleotide sequence represented by SEQ ID NO: 17 (TT>G);
R) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 18 (G>C);
S) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 19 (C>A);
T) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 20 (C>T);
U) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 21 (->CT);
V) a gene polymorphism (C> T) at the 301st base in the nucleotide sequence represented by SEQ ID NO: 22;
W) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 23 (A>T);
X) a gene polymorphism (G> A) at the 301st base in the base sequence represented by SEQ ID NO: 24;
Y) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 25 (->GA);
Z) a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 26 (AC>-); and
AA) gene polymorphism (T> G) at the 301st base in the base sequence represented by SEQ ID NO: 27;
(However, the parentheses indicate major allele> minor allele, and “-” indicates a base deletion.)
And (2) if the presence of a minor allele is found as a result of the test of (2) and (1), the subject is less sensitive to treatment with type I interferon A method of predicting a subject's sensitivity to treatment with type I interferon, comprising a step of determining;
[11] The prediction method according to [10] above, wherein the gene sample contains genomic DNA or RNA,
[12] The prediction method according to [10] or [11] above, wherein the subject is a patient with a disease to which treatment with type I interferon can be applied,
[13] The prediction method according to [12] above, wherein the patient with a disease to which treatment with type I interferon can be applied is an HCV-infected patient,
[14] The prediction method according to [13] above, wherein the HCV-infected patient is an HCV genotype 1b-infected patient, an HCV genotype 1a-infected patient, or an HCV genotype 2a-infected patient,
[15] The prediction method according to any one of [10] to [14], wherein the subject is an East Asian.
[16] (1) A gene polymorphism present in the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) gene regions located in human chromosome 19 short arm 19q13.13, using a gene sample derived from a subject, A step of testing a gene polymorphism at the 301st base in the nucleotide sequence represented by SEQ ID NO: 27, and (2) as a result of the test of (1), a homozygous gene polymorphism (TT) of a major allele A method for predicting the susceptibility of a subject to treatment with type I interferon, comprising the step of determining that the subject is highly sensitive to treatment with type I interferon, and [17] the subject is an East Asian The present invention relates to the prediction method according to [16].

According to the present invention, it is possible to provide a marker for predicting the therapeutic effect of a patient in IFN therapy, specifically, type I IFN therapy. That is, the present invention provides a genetic polymorphism on the gene region of IFNλ2 or IFNλ3, which is a marker for determining the therapeutic effect of IFN therapy in chronic hepatitis C. In addition, by predicting the genotype of a subject based on whether or not the polymorphism is present, it is possible to predict the therapeutic effect when the subject receives IFN therapy.
Since IFN therapy has been administered for a long period of more than half a year, and side effects have been reported (Adv. Intern. Med. 1994, 39, 241-275), if the therapeutic effect can be predicted in advance, It will be possible for the patient to select the optimal treatment method, determine the appropriate dose, and the duration of administration, which is beneficial for the patient.

It is a figure which shows the result of the genome-wide association analysis of SNPs related with the therapeutic effect of IFN therapy (Peginterferon-ribavirin combination therapy). The logarithm of the P value (Cochrane Armitage test) of each SNP was plotted against the position on the chromosome. The numbers on the horizontal axis indicate chromosome numbers. The peak of the SNP showing the minimum P value was located in the mapped region of the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) genes located on human chromosome 19 19q13.13. It is the figure which showed distribution of P value of genome-wide SNP related analysis by the quantile-quantile plot (Q-Qplot). The P value expected under the null hypothesis is plotted on the x-axis and the actual P value is plotted on the y-axis. The y = x line shows the distribution of P values under the null hypothesis. It is a figure which shows the reduction | decrease rate of the blood HCV-RNA 2 weeks after the start of IFN therapy (Peginterferon-ribavirin combination therapy) divided into the genotype of rs8099917 (T / G). The vertical axis is “blood HCV-RNA level (2W) 2 weeks after the start of IFN therapy (peginterferon / ribavirin combination therapy)” and “blood HCV-RNA level before IFN therapy (peginterferon / ribavirin combination therapy)” (0W) ”is a logarithmic conversion value with 10 as the base value divided by“ 0W ”, and represents the blood HCV-RNA decrease rate 2 weeks after the start of IFN therapy (Peginterferon / Ribavirin combination therapy). The dot indicates the value of each case, and the bar indicates the average value. The horizontal axis indicates the genotype. It is a figure which shows the decrease rate of blood HCV-RNA 4 weeks after the start of IFN therapy (peginterferon / ribavirin combination therapy), divided into rs8099917 (T / G) genotypes. The vertical axis is “blood HCV-RNA level (4W) 4 weeks after starting IFN therapy (peginterferon / ribavirin combination therapy)” and “blood HCV-RNA level before IFN therapy (peginterferon / ribavirin combination therapy)” (0W) ”is a logarithmic conversion value with 10 as the base value divided by“ 0W ”, and represents the blood HCV-RNA decrease rate 4 weeks after the start of IFN therapy (peginterferon-ribavirin combination therapy). The dot indicates the value of each case, and the bar indicates the average value. The horizontal axis indicates the genotype.

The present invention is described in detail below.
In this specification, IFNλ2 is
(A) a protein comprising the amino acid sequence represented by SEQ ID NO: 29 (Genbank Accession No .: NP_742150),
(B) a protein encoded by a polynucleotide comprising the base sequence represented by SEQ ID NO: 28 (Genbank Accession No .: NM — 172138),
(C) (c-1) in the amino acid sequence represented by SEQ ID NO: 29, wherein one or more amino acids are deleted, added, inserted or substituted, (c-2) amino acid sequence represented by SEQ ID NO: 29 An amino acid sequence having a sequence homology of 70% or more, or (c-3) a DNA having complementarity to a DNA having the 53rd to 655th nucleotide sequences of the nucleotide sequence represented by SEQ ID NO: 28; It is a protein consisting of an amino acid sequence encoded by DNA that hybridizes under stringent conditions, and having a binding activity to IL28Ra and IL10Rβ heteroreceptors.
In this specification, IFNλ3 is
(D) a protein comprising the amino acid sequence represented by SEQ ID NO: 31 (Genbank Accession No .: NP_742151),
(E) a protein encoded by a polynucleotide comprising the base sequence represented by SEQ ID NO: 30 (Genbank Accession No .: NM — 172139),
(F) (f-1) an amino acid sequence in which one or more amino acids are deleted, added, inserted or substituted in the amino acid sequence represented by SEQ ID NO: 31, (f-2) an amino acid sequence represented by SEQ ID NO: 31 An amino acid sequence having a sequence homology of 70% or more, or (f-3) a DNA having complementarity to a DNA having the 5th to 595th base sequences of the base sequence represented by SEQ ID NO: 30, It is a protein consisting of an amino acid sequence encoded by DNA that hybridizes under stringent conditions, and having a binding activity to IL28Ra and IL10Rβ heteroreceptors.
Here, “deletion, addition, insertion or substitution of amino acids” in the above (c-1) and (f-1) and “sequence homology of 70% or more in the above (c-2) and (f-2)” "Is a naturally occurring mutation caused by, for example, processing that the protein having the amino acid sequence shown in SEQ ID NO: 29 or 31 undergoes in the cell, species differences of individuals from which the protein is derived, individual differences, differences between tissues, etc. And artificial amino acid mutations.
Examples of the method for artificially performing the “amino acid deletion, addition or substitution” in the above (c-1) and (f-1) (hereinafter sometimes collectively referred to as amino acid modification) include, for example, Examples include a method of performing conventional site-directed mutagenesis on the DNA encoding the amino acid sequence represented by SEQ ID NO: 28 or 30 and then expressing this DNA by a conventional method. Examples of the site-directed mutagenesis method include a method using amber mutation (gapped duplex method, Nucleic Acids Res., 12, 9441-9456 (1984)), and a PCR method using a mutagenesis primer. Etc.
The number of amino acids modified as described above is at least one residue, specifically one or several, or more. The number of such modifications may be within a range in which the activity of IFNλ2 or IFNλ3 such as the binding activity of IL28Ra and IL10Rβ to the heteroreceptor of the protein can be found.
Of the deletions, additions or substitutions, the modification relating to amino acid substitution is particularly preferable. The substitution is more preferably substitution with an amino acid having similar properties such as hydrophobicity, charge, pK, and steric features. Such substitutions include, for example, i) glycine, alanine; ii) valine, isoleucine, leucine; iii) aspartic acid, glutamic acid, asparagine, glutamine; iv) serine, threonine; v) lysine, arginine; vi) phenylalanine, Substitution within the tyrosine group.

“Sequence homology” refers to sequence identity and similarity between two DNAs or two proteins. The “sequence homology” is determined by comparing two sequences that are optimally aligned over the region of the sequence to be compared. Here, the DNA or protein to be compared may have an addition or a deletion (for example, a gap) in the optimal alignment of the two sequences. Such sequence homology is calculated, for example, by creating an alignment using the ClustalW algorithm (Nucleic Acid Res., 22 (22): 4673-4680 (1994)) using Vector NTI. Can do. The sequence homology is measured using sequence analysis software, specifically Vector NTI, GENETYX-MAC or an analysis tool provided by a public database. The public database is generally available at, for example, a homepage address http://www.ddbj.nig.ac.jp.
The sequence homology in the present invention may be 70% or more, preferably 80% or more, more preferably 90% or more, and still more preferably 95% or more.

Regarding the “hybridize under stringent conditions” in the above (c-3) and (f-3), the hybridization used here is, for example, Sambrook J., Frisch EF, Maniatis T., Molecular. It can be carried out according to the usual methods described in the 2nd edition of cloning (Molecular Cloning 2nd edition), Cold Spring Harbor Laboratory press (Cold Spring Harbor Laboratory press) and the like. “Stringent conditions” means, for example, 6 × SSC (a solution containing 1.5M NaCl and 0.15M trisodium citrate is 10 × SSC), and a solution containing 50% formamide at 45 ° C. And the like (Molecular Biology, John Wiley & Sons, NY (1989), 6.3.1-6.3.6). The salt concentration in the washing step can be selected, for example, from 2 × SSC at 50 ° C. (low stringency conditions) to 0.2 × SSC at 50 ° C. (high stringency conditions). The temperature in the washing step can be selected, for example, from room temperature (low stringency conditions) to 65 ° C. (high stringency conditions). It is also possible to change both the salt concentration and the temperature.
In this specification, IFNλ2 corresponds to the above (a) to (c) and is not particularly limited as long as it maintains the physiological activity as IFNλ2. Here, the physiological activity of IFNλ2 includes binding activity to IL28Ra and IL10Rβ heteroreceptors and antiviral activity. The binding activity of IL28Ra and IL10Rβ to the heteroreceptor can be measured by the method described in Non-Patent Document 9. The antiviral activity can be measured by a method well known to those skilled in the art, such as the method described in Non-Patent Document 8.

Specific examples of the above-mentioned IFNλ2 (protein) include human IFNλ2 consisting of the amino acid sequence represented by SEQ ID NO: 29, mouse (Genbank Acc. No .: NP_001019844), rat (Genbank Acc. No .: XP_001065467), and the like. Mention orthologs from other animal species.
In this specification, the IFNλ2 gene represents a gene having a base sequence encoding the IFNλ2. Specifically, in addition to human IFNλ2 cDNA consisting of the base sequence represented by SEQ ID NO: 28, other animal species such as mouse (Genbank Acc. No .: NM_001024673), rat (Genbank Acc. No .: XM_001065467), etc. An ortholog can be mentioned.

Specific examples of the above-mentioned IFNλ3 (protein) include human IFNλ3 consisting of the amino acid sequence represented by SEQ ID NO: 31, chimpanzee (Genbank Acc. No .: XP_001135462), mouse (Genbank Acc. No .: NP_796370), chicken ( Genbank Acc. No .: NP_001121968), dogs (Genbank Acc. No .: XP_855366.), And other orthologs of other animal species can be mentioned.
In this specification, the IFNλ3 gene represents a gene having a base sequence encoding the IFNλ3. Specifically, in addition to human IFNλ3 cDNA consisting of the base sequence shown in SEQ ID NO: 30, chimpanzee (Genbank Acc. No .: XM_001135462), mouse (Genbank Acc. No .: NM_177396), chicken (Genbank Acc. No. : NM_001128496), dogs (Genbank Acc. No .: XM_850273), and other orthologs of other animal species.

I. Marker polymorphism and detection reagent thereof of the present invention The first aspect of the present invention relates to a reagent for predicting susceptibility to treatment with type I IFN described in [1] to [9] above. That is, the gene polymorphisms (SNP or DIP (deletion / insertion polymorphism)) shown in A) to Z) of [1] above are all polymorphisms that cause linkage disequilibrium with each other. Useful as a polymorphism. Here, the “linkage disequilibrium” is a polymorphism in a chain disequilibrium state in which the linkage disequilibrium coefficient r ² is 0.64 or more, preferably 0.8 or more. “Linkage disequilibrium coefficient r ² ” is defined as (A, a) for all polymorphisms and (B, b) for alleles of the second polymorphism. If the frequencies of the two haplotypes (AB, Ab, aB, ab) are P _AB , P _Ab , P _aB , P _ab , the following formula is obtained.
r ² = (P _AB P _ab −P _Ab P _aB ) ² / (P _AB + P _aB ) (P _aB + P _ab ) (P _AB + P _Ab ) (P _Ab + P _ab )
In addition, other known polymorphisms that are in linkage disequilibrium with the gene polymorphism can be identified using, for example, the HapMap database (http://www.hapmap.org/index.html.ja). As used herein, “(gene) polymorphism” is a change in one or more bases (substitution, deletion, insertion, transposition, inversion, etc.) on genomic DNA, and the change is 1 in the population. % Present at a frequency of more than%, for example, one base replaced with another base (SNP), one to several tens of bases deleted or inserted (DIP), 2 to several The number of repeats is different at the site where a sequence with 10 bases as one unit repeats (a microsatellite polymorphism with a repeat unit of 2-4 bases, a variable number of tandem repeat ( VNTR)) and the like, and SNP or DIP is preferred.

Here, “the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) gene region located in human chromosome 19 short arm 19q13.13” is a gene encoding human IFNλ2 represented by SEQ ID NO: 28, and SEQ ID NO: A genomic region containing a gene encoding human IFNλ3 represented by 30; in NCBI genome sequence ID number: NT_011109.15, a region represented by contig number 11987420 to 12029980 and a region about 5 kbp upstream and downstream thereof it can. The gene sequence of the gene region includes naturally occurring mutations due to individual differences, differences between tissues, and the like.
“Polynucleotide” is a concept that includes both RNA and DNA. Here, “DNA” includes not only double-stranded DNA but also each single-stranded DNA such as a sense strand (also referred to as a normal strand) and an antisense strand (also referred to as a complementary strand or reverse strand) constituting the DNA. Used for purposes.
Here, the antisense strand (complementary strand, reverse strand) means a polynucleotide having a base-complementary relationship based on the base pair relationship such as A: T and G: C with respect to the normal strand. Is.
The gene or DNA does not ask for distinction of the functional region, and can include, for example, an expression control region, a coding region, an exon, or an intron.
In addition, “RNA” is used to include not only single-stranded RNA but also single-stranded RNA having a complementary sequence thereto, and further double-stranded RNA composed thereof.
Furthermore, DNA and RNA chimeric molecules and DNA: RNA hybrids are also encompassed by the polynucleotides herein.

An “allele” is defined as one type when there are multiple genetic components that can occupy the same locus (locus) on a chromosome.
It is not a concept that necessarily includes a gene as a functional unit, and there are cases where multiple bases (sequences) exist at the same locus for gene polymorphisms such as SNP. That is, it shows a plurality of types of genes and bases located at homologous loci in a pair of homologous chromosomes derived from parents, and in this specification, gene polymorphisms such as SNPs located at homologous loci. For example, in the case of A for a chromosome derived from a father and G for a chromosome derived from a mother at a certain locus, the concepts of A allele and G allele are shown. English notation is A allele, G allele.
In the present specification, “major allele” means a base (sequence) that appears at a high frequency when a gene polymorphism exists at a corresponding gene locus. For example, assuming a SNP for A in the chromosome derived from the father and G in the chromosome derived from the mother, the frequency of the A allele is 80% and the frequency of the G allele is 20%. SNPs are called major alleles. On the other hand, when a genetic polymorphism is present at a corresponding gene locus, the base (sequence) that appears less frequently (in the above example, the SNP of G) is referred to as a minor allele.

In the present invention, the marker polymorphism described in A) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 1, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs955155. The polymorphism is represented by C or T (C>T; major allele> minor allele) in the genomic sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes, and the same applies to the following. ) Genetic polymorphism.
In the present invention, the marker polymorphism described in B) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 2, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs958039. The polymorphism is a gene polymorphism in which the base corresponding to contig number 11998519 is T or A (T> A) in the genomic sequence NT_011109.15 containing the human IFNλ2 and human IFNλ3 gene regions.
In the present invention, the marker polymorphism described in C) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 3, and NCBI [http: //www.ncbi.nlm ID number: rs35790907 in SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] provided by .nih.gov /]. The polymorphism is a genetic polymorphism in which the base corresponding to contig number 11998973 is A or T (A> T) in the genome sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in D) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 4. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12000930 is A or G (A> G) in the genome sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in E) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 5. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12000970 is G or A (G> A) in the genome sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in F) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 6. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12000122 is T or C (T> C) in the genome sequence NT_011109.15 including the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in [1] G) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 7, and NCBI [http: //www.ncbi.nlm ID number: rs8105790 in SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] provided by .nih.gov /]. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12000719 is T or C (T> C) in the genome sequence NT_011109.15 containing the human IFNλ2 and human IFNλ3 gene regions.
In the present invention, the marker polymorphism described in H) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 8. The polymorphism is a gene polymorphism in which the base corresponding to the contig number 12001341 is C or T (C> T) in the genome sequence NT_011109.15 including the human IFNλ2 and human IFNλ3 gene regions.
In the present invention, the marker polymorphism described in I) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 9, and NCBI [http: //www.ncbi.nlm ID number: rs4803217 in SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] provided by .nih.gov /]. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12002438 is C or A (C> A) in the genomic sequence NT_011109.15 including the human IFNλ2 and human IFNλ3 gene regions.
In the present invention, the marker polymorphism described in J) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 10, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs11881222. The polymorphism is a gene polymorphism in which the base corresponding to the contig number 12003141 is A or G (A> G) in the genomic sequence NT_011109.15 including the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in [1] K) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 11, and NCBI [http: //www.ncbi.nlm ID number: rs28416813 in SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] provided by .nih.gov /]. The polymorphism is a gene polymorphism in which the base corresponding to the contig number 12003862 is C or G (C> G) in the genomic sequence NT_011109.15 including the human IFNλ2 and human IFNλ3 gene regions.
In the present invention, the marker polymorphism described in L) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 12. The polymorphism is a genetic polymorphism in which the base corresponding to contig number 12004137 is C or T (C> T) in the genomic sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in [1] M) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 13, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs8107030. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12004937 is A or G (A> G) in the genomic sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in N) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 14, and NCBI [http: //www.ncbi.nlm ID number: rs73930703 in SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] provided by .nih.gov /]. The polymorphism is a gene polymorphism in which the base corresponding to the contig number 12005731 is C or T (C> T) in the genomic sequence NT_011109.15 containing the human IFNλ2 and human IFNλ3 gene regions.
In the present invention, the marker polymorphism described in [1] O) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 15, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs11882871. The polymorphism is a genetic polymorphism in which the base corresponding to contig number 12005828 is A or G (A> G) in the genomic sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in [1] P) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 16, and NCBI [http: //www.ncbi.nlm ID number: rs12971396 in SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] provided by .nih.gov /]. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12006084 is C or G (C> G) in the genomic sequence NT_011109.15 including the human IFNλ2 and human IFNλ3 gene regions.
In the present invention, the marker polymorphism described in Q) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 17. The polymorphism is the base sequence corresponding to contig numbers 12007372 and 12007373 in the genomic sequence NT_011109.15 containing the human IFNλ2 and human IFNλ3 gene regions, or G (single base deletion) (TT> G) It is a genetic polymorphism.
In the present invention, the marker polymorphism described in [1] R) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 18, and NCBI [http: //www.ncbi.nlm ID number: rs4803222 in SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] provided by .nih.gov /]. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12007571 is G or C (G> C) in the genome sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in S) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 19. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12008893 is C or A (C> A) in the genome sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in [1] T) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 20. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12009683 is C or T (C> T) in the genome sequence NT_011109.15 including the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in [1] U) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 21, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs10642535. The polymorphism is a gene polymorphism in which a CT is inserted between the bases corresponding to contig numbers 12010902 and 12010903 in the genomic sequence NT_011109.15 containing the human IFNλ2 and human IFNλ3 gene regions (-> CT).
In the present invention, the marker polymorphism described in [1] V) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 22, and NCBI [http: //www.ncbi.nlm ID number: rs8109889 in SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] provided by .nih.gov /]. The polymorphism is a gene polymorphism in which the base corresponding to the contig number 12010988 is C or T (C> T) in the genomic sequence NT_011109.15 including the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in [1] W) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 23, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs8113007. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12011321 is A or T (A> T) in the genomic sequence NT_011109.15 containing the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in [1] X) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 24, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs7248668. The polymorphism is a gene polymorphism in which the base corresponding to contig number 12012039 is G or A (G> A) in the genomic sequence NT_011109.15 including the regions of human IFNλ2 and human IFNλ3 genes.
In the present invention, the marker polymorphism described in Y) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 25. The polymorphism is a gene polymorphism in which GA is inserted between the bases corresponding to contig numbers 12012320 and 12012321 in the genomic sequence NT_011109.15 containing the human IFNλ2 and human IFNλ3 gene regions (-> GA).
In the present invention, the marker polymorphism described in Z) of [1] is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 26, and NCBI [http: //www.ncbi.nlm .nih.gov /] SNP database provided by dbSNP [http://www.ncbi.nlm.nih.gov/SNP/] ID number: SNP indicated by rs10612351. The polymorphism is a gene polymorphism in which the base corresponding to contig numbers 12013025 and 12013026 is AC or-: (no base) in the genomic sequence NT_011109.15 containing the human IFNλ2 and human IFNλ3 gene regions ( AC>-).
In the present invention, the marker polymorphism described in [10] AA) is a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 27, and NCBI [http: //www.ncbi.nlm. SNP database provided by nih.gov/] The SNP indicated by ID number: rs8099917 in dbSNP [http://www.ncbi.nlm.nih.gov/SNP/]. The polymorphism is a gene polymorphism in which the base corresponding to the contig number 12011383 is T or G (T> G) in the genomic sequence NT_011109.15 including the human IFNλ2 and human IFNλ3 gene regions.

The polynucleotide capable of detecting the marker polymorphism of the present invention is useful for predicting whether or not a subject is sensitive to treatment with type I IFN. As a result of disease control correlation analysis, it was shown that the frequency of minor alleles in the marker polymorphisms of the present invention was significantly higher in the non-effective group than in the effective group treated with type I IFN. Therefore, by testing whether a subject has a minor allele in the marker polymorphism of the present invention, it can be predicted whether the subject is sensitive to treatment with type I IFN. Accordingly, the reagent for predicting susceptibility to treatment with type I IFN of the present invention contains a polynucleotide capable of detecting at least a minor allele in the marker polymorphism of the present invention. For example, in the marker polymorphism of A) in [1] above, it is a continuous 10-200 base sequence containing the 301st base in the base sequence represented by SEQ ID NO: 1 or its complementary strand sequence. Thus, the presence of a sequence whose base is T (A in the case of a complementary strand sequence) can be detected.
Specifically, the polynucleotide capable of detecting the marker polymorphism of the present invention is a gene chip such as Taqman probe method, Invader probe method (Third Wave), GENECHIP SNP ARRAY (AFFYMETRIX) or Beadchip (ILLUMINA). It is used as a primer or probe in known gene analysis methods such as SNP typing method, direct sequencing method, SSCP method (single-stranded conformational polymorphism analysis), ASP-PCR method (allele-specific primer PCR analysis) and the like. That is, a primer is a pair of polynucleotides that can amplify a human IFNλ2 or IFNλ3 gene fragment containing the marker polymorphic site of the present invention, and a probe is a human IFNλ2 or IFNλ3 gene containing the polymorphic site. A polynucleotide that can hybridize to a region under stringent conditions. Here, “stringent conditions” have the same meaning as described above.
The length of the polynucleotide capable of detecting the marker polymorphism of the present invention is not particularly limited as long as it can detect human IFNλ2 or IFNλ3 gene fragment having 10 to 200 consecutive base sequences including the polymorphic site. Specifically, the length can be appropriately selected and set according to the use of the polynucleotide.
When the polynucleotide of the present invention is used as a primer, one having a base length of 10 bp to 200 bp, preferably 15 bp to 50 bp, more preferably 15 bp to 35 bp can be exemplified. When used as a detection probe, those having a base number of 10 bp to 200 bp, preferably 10 bp to 50 bp, more preferably 15 bp to 30 bp can be exemplified.

  In the marker polymorphisms of the present invention, the minor allele is an insensitive allele for treatment with type I IFN, while the major allele is a susceptible allele. That is, when a subject has a genotype that is homozygous for a major allele, the frequency of sensitivity to treatment with type I IFN is significantly greater than for heterozygous or minor alleles. high. Therefore, it is desirable that the reagent for predicting susceptibility to treatment with type I IFN of the present invention further contains a polynucleotide capable of detecting a major allele in the marker polymorphism of the present invention.

When a polynucleotide capable of detecting the marker polymorphism of the present invention is used as a probe, the probe may contain an additional sequence (a sequence not complementary to genomic DNA) suitable for detecting the polymorphism. For example, the allele probe used in the Invader probe method has an additional sequence called a flap at the 5 ′ end of the base at the polymorphic site.
The probe may be a suitable labeling agent such as, for example, a radioisotope (eg, ¹²⁵ I, ¹³¹ I, ³ H, ¹⁴ C, etc.), an enzyme (eg, β-galactosidase, β-glucosidase, alkaline phosphatase, Peroxidase, malate dehydrogenase, etc.), fluorescent substances (eg, fluorescamine, fluorescein isothiocyanate, etc.), luminescent substances (eg, luminol, luminol derivatives, luciferin, lucigenin, etc.) . Alternatively, a quencher (quenching substance) that absorbs fluorescence energy emitted by the fluorescent substance may be further bound in the vicinity of the fluorescent substance (eg, FAM, VIC, etc.). In such an embodiment, the fluorescence is detected by separating the fluorescent substance and the quencher during the detection reaction.

When a polynucleotide capable of detecting the marker polymorphism of the present invention is used as a primer, the primer contains an additional sequence suitable for detection of the polymorphism (a sequence not complementary to genomic DNA), for example, a linker sequence. May be.
In addition, the primer may be an appropriate labeling agent such as, for example, a radioisotope (eg, ¹²⁵ I, ¹³¹ I, ³ H, ¹⁴ C, etc.), an enzyme (eg, β-galactosidase, β-glucosidase, alkaline phosphatase, Peroxidase, malate dehydrogenase, etc.), fluorescent substances (eg, fluorescamine, fluorescein isothiocyanate, etc.), luminescent substances (eg, luminol, luminol derivatives, luciferin, lucigenin, etc.) .

The nucleic acid probes and / or primers are each separately (or mixed if possible) in water or in an appropriate buffer (eg, TE buffer) at an appropriate concentration (eg, 2 × to 20 ×). And can be stored at about -20 ° C.
The reagent for predicting susceptibility to treatment with type I IFN of the present invention may further contain other components necessary for carrying out the method as a component, depending on the polymorphism detection method. For example, when the reagent is for polymorphism detection by TaqMan PCR method, the reagent includes 10 × PCR reaction buffer, 10 × MgCl ₂ aqueous solution, 10 × dNTPs aqueous solution, Taq DNA polymerase (5 U / μL), etc. Can further be included.

II. Method for Predicting Sensitivity to Treatment with Type I IFN The present invention also predicts whether or not a subject has sensitivity to treatment with type I IFN by examining the genotype of the subject, [10] to [17] The prediction method as described above is included.

Here, the subject-derived gene sample to be measured is not particularly limited as long as it is a biological sample containing the gene of the subject (patient, etc.), that is, genomic DNA and RNA, and is appropriately selected according to the type of detection method used. (However, when RNA or a polynucleotide derived therefrom (eg, cDNA) is used as a gene sample, the marker polymorphism to be detected is selected from those present in the exon portion of the human IFNλ2 or IFNλ3 gene. ) The sample may be, for example, a biological tissue of a subject, specifically, blood, liver biopsy, buccal mucosa and the like, and genomic DNA or total RNA prepared therefrom according to a conventional method may be used. Various polynucleotides prepared based on RNA may be used. Any method known to those skilled in the art may be used to prepare genomic DNA and RNA from a subject-derived sample.
The race of the subject is not particularly limited, but is preferably East Asian, and more preferably Japanese.

Specifically, the above-described polynucleotide capable of detecting the marker polymorphism of the present invention is used as a primer or a probe according to a conventional method, Taqman probe method, Invader probe method (Third Wave Technologies), GENECHIP SNP ARRAY (AFFYMETRIX And SNP typing method using gene chips such as Beadchip (ILLUMINA), direct sequencing method, SSCP method (single-stranded conformational polymorphism analysis), ASP-PCR method (allele-specific primer PCR analysis), etc. What is necessary is just to test by the well-known method of detecting a gene specifically.
For example, using the Taqman probe method and the Invader probe method, the following groups:
A) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 1;
B) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 2 (T>A);
C) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 3 (A>T);
D) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 4 (A>G);
E) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 5 (G>A);
F) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 6 (T>C);
G) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 7 (T>C);
H) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 8 (C>T);
I) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 9 (C>A);
J) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 10 (A>G);
K) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 11 (C>G);
L) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 12;
M) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 13 (A>G);
N) a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 14 (C>T);
O) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 15 (A>G);
P) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 16 (C>G);
Q) Gene polymorphism at the 301st base in the nucleotide sequence represented by SEQ ID NO: 17 (TT>G);
R) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 18 (G>C);
S) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 19 (C>A);
T) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 20 (C>T);
U) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 21 (->CT);
V) a gene polymorphism (C> T) at the 301st base in the nucleotide sequence represented by SEQ ID NO: 22;
W) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 23 (A>T);
X) a gene polymorphism (G> A) at the 301st base in the base sequence represented by SEQ ID NO: 24;
Y) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 25 (->GA);
Z) a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 26 (AC>-); and
AA) gene polymorphism (T> G) at the 301st base in the base sequence represented by SEQ ID NO: 27;
(However, the parentheses indicate major allele> minor allele, and “-” indicates a base deletion.)
It can be confirmed that the human IFNλ2 or IFNλ3 gene fragment containing the polymorphic site of the gene polymorphism selected from is specifically generated. Methods for predicting SNP genotypes using the Taqman probe method are well known to those skilled in the art. For example, PCR primers and SNPs that detect SNP regions using TaqMan (registered trademark) SNP Genotyping Assays reagents commercially available from Applied Biosystems. Fluorescently labeled TaqMan probes that identify alleles and genomic DNA from the subject's genetic sample are mixed, reacted according to the package insert, and analyzed for fluorescence signals to analyze the SNP alleles contained in the sample. A method for detecting the genotype can be exemplified (Nat Genet 2003; 34: 395-402).
In addition, a method for identifying an SNP genotype using an Invader probe is well known to those skilled in the art, and an Invader probe and a reaction reagent for identifying an SNP allele marketed by Third Wave Technologies are mixed to derive from a subject's gene sample. A method for detecting the genotype of an SNP allele contained in a sample can be exemplified by reacting genomic DNA as a template according to the method recommended by kit and detecting a fluorescent signal with a general-purpose fluorescent plate reader (J Hum Genet 2001; 46: 471-477).

When the Northern blot method is used, a polynucleotide that can detect the marker polymorphism described in [1] above may be used as a probe. Specifically, the probe radioisotope ⁽³² P, ³³ P, etc.: RI) labeled with a, fluorescent substance, it was nylon membrane or the like is RNA hybridized from transfer cells to the usual manner Thereafter, the formed double strand of the probe (DNA or RNA) and RNA is used as a signal derived from the probe label (RI or fluorescent substance) as a radiation detector (BAS-1800II, manufactured by Fuji Film) Or the method of detecting and measuring with a fluorescence detector can be illustrated. Further, using the AlkPhos Direct Labeling and Detection System (manufactured by Amersham Pharamcia Biotech), the probe is labeled in accordance with the protocol, hybridized with cell-derived RNA, and then the signal derived from the labeled product of the probe is multibiotic. A method of detecting and measuring with Major STORM860 (manufactured by Amersham Pharmacia Biotech) can also be used.

The genotype of the subject can also be detected using a gene chip such as GENECHIP SNP ARRAY (AFFYMETRIX) or Beadchip (ILLUMINA), and includes a polynucleotide sequence that can detect the marker polymorphism of the present invention. A polynucleotide of any length up to 10-100 bp can be used as a probe. Methods using GENECHIP SNP ARRAY (AFFYMETRIX) and Beadchip (ILLUMINA) are well known to those skilled in the art, and kits from AFFYMETRIX and ILLUMINA
According to a recommended method, a method of detecting a genotype of an SNP allele contained in a sample by hybridizing a DNA chip containing the polynucleotide of the present invention in a probe with a gene sample derived from a subject. it can.

“Type I IFN” includes IFN-α and IFN-β. Here, IFN-α is further classified into small isoforms with slightly different specificities, but IFN-α in this specification is a mixture of these isoforms, and natural IFN-α The concept encompasses all recombinant proteins from which these isoforms have been isolated. Examples of isoforms include IFN-α2a and IFN-α2b.
“High sensitivity to treatment with type I IFN” means a state exhibiting the expected pharmacological action when administered type I IFN. Specifically, if a patient suffers from a viral infection, the blood ALT (alanine transaminase) level, which is an indicator of decreased blood viral RNA levels, complete disappearance, and improved liver function Normalization is mentioned.
Patients who are highly sensitive to treatment with type I IFN can be expected to benefit from treatment with type I IFN.
Diseases for which treatment with type I IFN is effective, that is, diseases to which treatment for type I IFN can be applied, include HCV, HBV (hepatitis B virus), and other subacute sclerosis Examples include encephalitis, HTLV-I (human adult T cell leukemia virus-I) myelopathy, and carcinogenesis from cirrhosis. Also included are renal cancer, multiple myeloma, leukemia and the like.
Examples of patients with diseases to which the treatment of type I IFN can be applied include patients suffering from the above-mentioned diseases. When the patient is an HCV-infected patient, the HCV genotype 1b patient is more sensitive to treatment with type I IFN and is more significantly correlated with the marker polymorphism of the present invention. It is preferable to carry out.
On the other hand, in patients with low sensitivity to treatment with type I IFN, the amount of blood viral RNA does not decrease even after administration of type I IFN, it does not disappear completely even if it decreases, blood ALT levels do not normalize once Even if it disappears, it shows signs such as viral RNA re-growth after treatment.

  When a subject has a minor allele as a result of testing for at least one genetic polymorphism of A) to Z) and AA) described above in a subject-derived gene sample, the subject is susceptible to treatment with type I IFN. Can be predicted to be low. More preferably, as a result of testing for at least one gene polymorphism of A) to Z) and AA) described above, when the major allele is homozygous, it is highly susceptible to treatment with type I IFN and is heterozygous. In this case, the sensitivity to treatment with type I IFN is lower than when the major allele is homozygous, and the sensitivity to treatment with type I IFN is even lower when the minor allele is homozygous.

  EXAMPLES Hereinafter, although an Example demonstrates this invention in more detail, this invention is not limited to these Examples.

[SNP genotype determination in patients with chronic hepatitis C] (case control group 1 (screening study))
A case-control correlation analysis (case-control group 1 (screening study)) was performed to analyze the relationship between the therapeutic effects of IFN therapy (Pegylated interferon / ribavirin combination therapy) and SNPs.
Infected with HCV genotype 1b after completing IFN therapy (Peginterferon and ribavirin combination therapy) for chronic hepatitis C at Hiroshima University Hospital, Hiroshima University Hospital, and Toranomon Hospital, and consent to participate in this study Genomic DNA was prepared from the peripheral blood of 594 patients, and the SNP genotype was determined using Human610-QuadBeadChip (Illumina). Human610-Quad BeadChip (Illumina) is a gene chip in which more than 550,000 tag SNPs derived from HapMap data covering the genome at high density are evenly arranged. All 594 patients infected with HCV genotype 1b are Japanese. Nine cases were judged to be closely related, and two cases were excluded from the analysis due to the high error rate, and 583 subjects were included in the analysis (see Table 1: Case Control Group 1 (Screening Study)). Unrelated persons were confirmed using PLINK software. The presence or absence of population stratification was analyzed with EIGENSTRAT.
For the case control group 1 (screening study), an inflation factor was calculated from the P value obtained as a result of the association analysis of the case control group 1 (screening study) using the genomic control method. Cases with a call rate of less than 98%, SNPs with a call rate of less than 99%, non-autosomal SNPs, MAF <0.01% SNPs, SNPs not in Hardy-Weinberg equilibrium (p <1 × 10 ^-6 ) were excluded from the analysis. .

Based on the results of genotyping of about 500,000 SNPs using Human610-QuadBeadChip (Illumina), we analyzed the relationship between the therapeutic effects of IFN therapy (Peginterferon / Ribavirin combination therapy) and SNPs.
Association analysis with IFN therapy (Peginterferon and ribavirin combination therapy) was performed using the Cochrane Armitage test, and Chi-square test was also used as a reference for the allele model, allele dominant inheritance model, and allele recessive inheritance model. . The logarithmically transformed values of the obtained P values were plotted on the chromosome. The peak of SNPs showing the minimum P value was located in the mapped region of the interferon λ2 (IFNλ2) and / or interferon λ3 (IFNλ3) gene located in human chromosome 19 19q13.13 (FIG. 1). See).
The SNP showing the minimum P value is the SNP database dbSNP [http://www.ncbi.nlm.nih.gov/SNP provided by NCBI [http://www.ncbi.nlm.nih.gov/]. ID number in /]: SNP indicated by rs8099917.
The P value in the Cochrane Armitage test for rs8099917 was 6.5 × 10 ⁻⁸ . The results are shown in Table 2 below.
The polymorphism was a gene polymorphism at the base represented by contig number 12011383 in the genomic sequence NT_011109.15 containing the human IFNλ2 and human IFNλ3 gene regions. In genome sequence NT_011109.15, contig number 12011383 was located upstream of the region of interferon λ3 (IFNλ3) gene.
Linkage disequilibrium mapping in the mapped region of the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) genes located on human chromosome 19q13.13 is shown in the HapMap database (http://www.hapmap.org/cgi -Perl / gbrowse / hapmap27_B36 /) "Data source: HapMap Data Rel 27 Phase II + III, Feb09, on NCBI B36 assembly, db SNP b126" The linkage disequilibrium block was found to be a region represented by contig numbers 11987642 to 12029972 in NCBI genome sequence ID number NT_011109.15.

[Identification of rs8099917 associated with IFN treatment effect]
In Example 1, the allelic frequency and genotype frequency of the identified rs8099917 were analyzed. The identified rs8099917 matches the registered allele in NCBI, and is hereinafter referred to as rs8099917 (T / G) (major allele / minor allele).
Table 2 shows the analysis results of the identified rs8099917 (T / G) allele model, that is, the allele frequency.
In case-control group 1 (screening study) (Table 1) who are HCV genotype 1b-infected patients, a 2 × 2 contingency table of therapeutic effect (effective / non-effective) and SNP allele (T / G) was prepared. P-value (two-sided P-value by chi-square test), odds ratio and 95% confidence interval were calculated.
As a result, in patients with HCV genotype 1b infection, the frequency of the T allele was 90.0% for the effective group, 78.5% for the non-effective group, and the frequency for the G allele was 21.5% for the effective group 10.0%. There was a significant difference between the two groups (P value 6.5 X 10 ^-8 , odds ratio 2.46, 95% confidence interval (1.76-3.43)).
From the result of analyzing the allele frequency of the identified rs8099917 (T / G),
(a) If you have a T allele, you are more sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than if you have a G allele, or
(b) When the G allele is present, the sensitivity to treatment with IFN therapy (peginterferon / ribavirin combination therapy) is lower than when the T allele is present.
It was found that it can be predicted.
Table 2 similarly shows the results of analyzing the association between the effective group and the non-effective group regarding the genotype frequency of rs8099917 (T / G).
In patients with HCV genotype 1b infection, the frequency of GG homozygous type was 3.6% for the non-effective group compared to 1.3% for the effective group, and the frequency for the TG-zygous type was 17.4% for the effective group It was 35.8%. The frequency of the TT homozygous type was 60.6% for the non-effective group compared to 81.3% for the effective group.
In the G allele dominant inheritance model, the phenotype of TG heterozygous patients was classified as G allele traits, assuming that the traits of the G allele were dominant over the traits of the T allele. Create a 2 × 2 contingency table of therapeutic effect (effective / non-effective) and genotype (TT / TG + GG), and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated. As a result, the P value was 3.5 × 10 ⁻⁸ , and a significant difference was observed between the two groups.
In the G-allelic recessive inheritance model, the T / G heterozygous patient phenotype was classified as a T allele trait, assuming that the T allele trait was dominant over the G allele trait. Create a 2 × 2 contingency table of therapeutic effect (superior / non-responsible) and genotype (TT + TG / GG) and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated. As a result, the P value was 0.10,
From the above results,
(a) When the T allele is homozygous, it is more sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than when the G allele is homozygous or heterozygous. Or
(b) When the G allele is homozygous or heterozygous, it is less sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than when the T allele is homozygous. ,
It was found that it can be predicted.

[Determination of SNP genotype in patients with chronic hepatitis C] (Case Control Group 2 (Replication Study 1))
Table 1 shows the results of an analysis of the relationship between the therapeutic effect of IFN therapy (peginterferon and ribavirin combination therapy) on HCV genotype 1b infection and the allele and genotype frequencies of rs8099917 (T / G). For the case control group 2 shown in (Replication Study 1), the genotype of rs8099917 (T / G) was determined.
Infected with HCV genotype 1b after completing IFN therapy (Peginterferon and ribavirin combination therapy) for chronic hepatitis C at Hiroshima University Hospital, Hiroshima University Hospital, and Toranomon Hospital, and consent to participate in this study Genomic DNA was prepared from the peripheral blood of 185 patients (case control group 2 (Replication Study 1) and genotype of rs8099917 was determined. Taqman method (Nat Genet 2003; 34: 395-402) The method followed the method of each cited document.
The allelic frequency and genotype frequency of rs8099917 (T / G) were analyzed for HCV genotype 1b-infected patients (case control group 2 (Replication Study 1)).
2 × 2 contingency table of therapeutic effect (effective / non-effective) and SNP allele (T / G) frequency in HCV genotype 1b-infected patients shown in Table 1 (case control group 2 (Replication Study 1)) And calculated P-value (two-sided P-value by chi-square test), odds ratio and 95% confidence interval. The results are shown in Table 3 below.
As a result, in patients with HCV genotype 1b infection (case control group 2 (Replication Study 1)), the frequency of the T allele was 92.7% in the effective group and 82.7% in the non-effective group, and the frequency of the G allele was significant. There was a significant difference between the two groups, with 5.9% in the group and 17.3% in the ineffective group. (P value 1.9 X 10 ^-3 , odds ratio 3.31, 95% confidence interval (1.50-7.30))
From this result, by determining the allele frequency of rs8099917 (T / G) of patients infected with HCV genotype 1b,
(a) If you have a T allele, you are more sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than if you have a G allele, or
(b) When the G allele is present, the sensitivity to treatment with IFN therapy (peginterferon / ribavirin combination therapy) is lower than when the T allele is present.
It was found that it can be predicted.
Table 3 similarly shows the results of analyzing the relationship with the therapeutic effect for the genotype frequency of rs8099917 (T / G).
In HCV genotype 1b-infected patients (case control group 2 (replication study 1)), the frequency of GG homozygous type was 3.4% for non-responding group versus 0% for effective group (0 patients). The frequency was 11.9% in the effective group and 28.0% in the non-effective group. The frequency of the TT homozygous type was 88.6% in the effective group and 68.6% in the non-effective group.
In the G allele dominant inheritance model, the phenotype of TG heterozygous patients was classified as G allele traits, assuming that the traits of the G allele were dominant over the traits of the T allele. Create a 2 × 2 contingency table of therapeutic effect (effective / non-effective) and genotype (TT / TG + GG), and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated. As a result, the P value was 3.1 × 10 ⁻³ , and a significant difference was observed between the two groups.
In the G-allelic recessive inheritance model, the T / G heterozygous patient phenotype was classified as a T allele trait, assuming that the T allele trait was dominant over the G allele trait. Create a 2 × 2 contingency table of therapeutic effect (superior / non-responsible) and genotype (TT + TG / GG) and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated. As a result, a P value of 0.3 showed no significant association in this model.
From the above results, by determining the genotype frequency of rs8099917 (T / G) of patients infected with HCV genotype 1b,
(a) When the T allele is homozygous, it is more sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than when the G allele is homozygous or heterozygous. Or
(b) When the G allele is homozygous or heterozygous, it is less sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than when the T allele is homozygous. It was found that it can be predicted.

[Determination of SNP genotype in patients with chronic hepatitis C] (Case Control Group 3 (Replication Study 2))
To further examine the results of an analysis of the relationship between the therapeutic effects of IFN therapy (IFN-α monotherapy) and rs8099917 (T / G) allele frequency and genotype frequency for patients infected with HCV genotype 1b, Table 1 For the case control group 3 shown (Replication Study 2), genotype determination of rs8099917 (T / G) was performed.
I completed IFN therapy (IFN-α monotherapy) for chronic hepatitis C at Hiroshima University Hospital, Hiroshima University School of Medicine, and Toranomon Hospital, and was infected with HCV genotype 1b with consent to participate in this study Genomic DNA was prepared from 750 patients and the genotype of rs8099917 (T / G) was determined. The Taqman method (Nat Genet 2003; 34: 395-402) was used for the determination, and the method followed the method of each cited reference.
For HCV genotype 1b-infected patients (case control group 3 (Replication Study 2)), the allele frequency and genotype frequency of rs8099917 (T / G) were analyzed.
In Table 1, patients with HCV genotype 1b infection (case control group 3 (Replication Study 2)), 2 × 2 contingency table for therapeutic effect (effective / non-effective) and SNP allele (T / G) And calculated P-value (two-sided P-value by chi-square test), odds ratio and 95% confidence interval. The results are shown in Table 4.
As a result, in HCV genotype 1b-infected patients (case control group 3 (Replication Study 2)), the frequency of the T allele was 95.9% for the effective group, 86.1% for the non-effective group, and the frequency for the G allele was significant. There was a significant difference between the two groups, with 4.1% in the group and 13.9% in the non-effective group. (P value 4.1 X ^10-8 , odds ratio 3.76, 95% confidence interval (2.27-6.21))
From this result, by determining the allelic frequency of rs8099917 (T / G) in patients infected with HCV genotype 1b,
(a) If you have a T allele, you are more sensitive to treatment with IFN therapy (IFN-α monotherapy) than if you have a G allele, or
(b) If the G allele is present, the sensitivity to treatment with IFN therapy (IFN-α monotherapy) is lower than when the T allele is present.
It was found that it can be predicted.
Table 4 similarly shows the results of analyzing the association between the effective group and the non-effective group regarding the genotype frequency of rs8099917 (T / G).
In HCV genotype 1b-infected patients (case control group 3 (Replication Study 2)), the frequency of GG homozygous type was 1.9% for non-effective group compared to 0.5% for effective group, and the frequency of TG-zygous type was significant. The efficacy group was 7.4% and the non-response group was 24.2%. The frequency of TT homozygous type was 93.9% in the effective group and 73.9% in the non-effective group.
In the G allele dominant inheritance model, the phenotype of TG heterozygous patients was classified as G allele traits, assuming that the traits of the G allele were dominant over the traits of the T allele. Create a 2 × 2 contingency table of therapeutic effect (effective / non-effective) and genotype (TT / TG + GG), and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated. As a result, the P value was 2.4 × 10 ⁻⁸ , and a significant difference was observed between the two groups.
In the G-allelic recessive inheritance model, the T / G heterozygous patient phenotype was classified as a T allele trait, assuming that the T allele trait was dominant over the G allele trait. Create a 2 × 2 contingency table of therapeutic effect (superior / non-responsible) and genotype (TT + TG / GG) and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated. As a result, P value 0.19 showed no significant association in this model.
From the above results, by determining the genotype frequency of rs8099917 (T / G) of patients infected with HCV genotype 1b,
(a) When the T allele is homozygous, it is more sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than when the G allele is homozygous or heterozygous. Or
(b) When the G allele is homozygous or heterozygous, it is less sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than when the T allele is homozygous. ,
It was found that it can be predicted.

[Determination of SNP genotype in patients with chronic hepatitis C] (Case Control Group 4 (Replication Study 3))
To verify the relationship between the therapeutic effect of IFN therapy (IFN-α monotherapy) on HCV genotype 2a patients and the allelic and genotype frequencies of rs8099917 (T / G), the results are shown in Table 1. For case control group 4 (Replication Study 3), genotype determination of rs8099917 (T / G) was performed.
For case control group 4 (Replication Study 3), which shows the relationship between the therapeutic effect of IFN therapy (IFN-α monotherapy) and the IFN therapeutic effect of rs8099917 (T / G), rs8099917 (T / G) The genotype was determined and analyzed. Infected with HCV genotype 2a that completed IFN therapy (IFN-α monotherapy) for chronic hepatitis C at Hiroshima University Hospital, Hiroshima University Hospital, and Toranomon Hospital, and consented to participate in this study Genomic DNA was prepared from 513 patients and the genotype of rs8099917 (T / G) was determined. The Taqman method (Nat Genet 2003; 34: 395-402) was used for the determination, and the method followed the method of each cited reference.
The allelic and genotypic frequencies of rs8099917 (T / G) were analyzed for HCV genotype 2a infected patients (case control group 4 (Replication Study 3)).
In Table 1, patients with HCV genotype 2a infection (Case Control Group 4 (Replication Study 3)): 2 × 2 contingency table for therapeutic effect (effective / non-effective) and SNP allele (T / G) And calculated P-value (two-sided P-value by chi-square test), odds ratio and 95% confidence interval. The results are shown in Table 5.
As a result, in patients with HCV genotype 2a infection (case control group 4 (Replication Study 3)), the frequency of T allele was 91.9% in the effective group, 85.4% in the non-effective group, and the frequency of the G allele was effective. The non-effective group was 14.6% compared with 8.1% in the group, and there was a significant difference between the two groups. (P value 1.4 X 10 ^-3 , odds ratio 1.92, 95% confidence interval (1.28-2.88))
From this result, by determining the allele frequency of rs8099917 (T / G) of patients infected with HCV genotype 2a,
(a) If you have a T allele, you are more sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than if you have a G allele, or
(b) When the G allele is present, the sensitivity to treatment with IFN therapy (peginterferon / ribavirin combination therapy) is higher than when the T allele is present.
It was found that it can be predicted.
Table 5 similarly shows the results of analyzing the association between the effective group and the non-effective group regarding the genotype frequency of rs8099917 (T / G). In the G allele dominant inheritance model, the phenotype of TG heterozygous patients was classified as the G allele trait, assuming that the trait of the G allele was dominant over the trait of the T allele. Create a 2 × 2 contingency table of therapeutic effect (effective / non-effective) and genotype (TT / TG + GG), and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated.
In the G-allelic recessive inheritance model, the T / G heterozygous patient phenotype was classified as a T allele trait, assuming that the T allele trait was dominant over the G allele trait. A 2 × 2 contingency table of therapeutic effect (effective / non-effective) and genotype (TT + TG / GG) was prepared and analyzed in the same manner.
In patients with HCV genotype 2a infection, the frequency of GG homozygous type was 2.9% in the non-effective group compared to 0.9% in the effective group, and the frequency of TG-zygous type was in the non-effective group compared to 14.6% in the effective group It was 23.4%. The frequency of the TT homozygous type was 73.7% in the non-effective group compared to 84.5% in the effective group.
In the G allele dominant inheritance model, the phenotype of TG heterozygous patients was classified as G allele traits, assuming that the traits of the G allele were dominant over the traits of the T allele. Create a 2 × 2 contingency table of therapeutic effect (effective / non-effective) and genotype (TT / TG + GG), and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated. As a result, the P value was 1.4 × 10 ⁻³ and judged to be significant.
In the G-allelic recessive inheritance model, the T / G heterozygous patient phenotype was classified as a T allele trait, assuming that the T allele trait was dominant over the G allele trait. Create a 2 × 2 contingency table of therapeutic effect (superior / non-responsible) and genotype (TT + TG / GG) and calculate P value (two-sided P value by chi-square test), odds ratio, and 95% confidence interval Calculated. As a result, the P value was 0.12.
Based on the above results, by determining the genotype frequency of rs8099917 (T / G) in patients infected with HCV genotype 2a,
(a) When the T allele is homozygous, it is more sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than when the G allele is homozygous or heterozygous. Or
(b) When the G allele is homozygous or heterozygous, it is less sensitive to treatment with IFN therapy (peginterferon / ribavirin combination therapy) than when the T allele is homozygous. It was found that it can be predicted.

[SNP genotype determination in patients with chronic hepatitis C] (integration by meta-analysis)
The Mantel-Haenzel test was performed to integrally evaluate the analysis results of four cohorts (case control groups 1, 2, 3 and 4 (Table 1)). Table 6 shows the results.
In the allele model (T vs G), the integrated P value was 1.73 X 10 ^-18 , the integrated odds ratio was 2.63, and the 95% confidence interval (2.11-3.27).
From this result, in patients infected with HCV genotypes 1b and 2a, in IFN therapy (IFN-α monotherapy or peginterferon / ribavirin combination therapy), by determining the allele frequency of rs8099917 (T / G),
(a) If the T allele is present, it is 2.63 times more likely to be effective in treatment with IFN therapy (IFN-α monotherapy or peginterferon / ribavirin combination therapy) compared to the G allele Expensive or
(b) If the G allele is present, it may be less effective in treatment with IFN therapy (IFN-α monotherapy or peginterferon / ribavirin combination therapy) compared to the T allele. It was found that it can be predicted to be twice as high.

[Prediction of early viral reduction rate of IFN therapy (Pegylated interferon / ribavirin combination therapy)]
We examined whether rs8099917 (T / G) genotype was associated with the early response of IFN therapy (Pegylated interferon / ribavirin combination therapy). Infected with HCV genotype 1b after completing IFN therapy (Peginterferon and ribavirin combination therapy) for chronic hepatitis C at Hiroshima University Hospital, Hiroshima University Hospital, and Toranomon Hospital, and consent to participate in this study Genomic DNA was prepared from these patients and the genotype of rs8099917 (T / G) was determined.
The Taqman method (Nat Genet 2003; 34: 395-402) was used for the determination, and the method followed the method of each cited reference. The amount of HCV-RNA in blood was quantified for the following (1) to (3). Quantification of the amount of HCV-RNA was performed by the original method, the high range method, and the TaqMan method.
The quantifiable range of the amount of HCV-RNA in these techniques is 0.5-850 KIU / ml, 5-5000 KIU / ml, and 1.2-7.8 log IU, respectively.
The following (1) to (3):
(1) Blood HCV-RNA level before IFN therapy (Peginterferon / Ribavirin combination therapy) (2) Blood HCV-RNA level 2 weeks after IFN therapy (Peginterferon / Ribavirin combination therapy) (3) IFN therapy (Peginterferon / ribavirin combination therapy) The results of blood HCV-RNA levels 4 weeks after the start were shown in FIGS.

FIG. 3 shows the reduction rate of blood HCV-RNA 2 weeks after the start of IFN therapy (peginterferon / ribavirin combination therapy), divided into rs8099917 (T / G) genotypes. The number of patients analyzed was 513.
The horizontal axis is “blood HCV-RNA level 2 weeks after starting IFN therapy (peginterferon / ribavirin combination therapy)” divided by “blood HCV-RNA level before IFN therapy (peginterferon / ribavirin combination therapy)”. The logarithmic conversion value with the base 10 as the base value, and represents the rate of decrease in blood HCV-RNA 2 weeks after the start of IFN therapy (Peginterferon-ribavirin combination therapy). The vertical axis shows the rs8099917 (T / G) genotype in order from the top: (a) GG homozygous type (GG), (b) GT heterozygous type (GT), and (c) TT homozygous type (TT ). When the correlation coefficient between the genotype of rs8099917 (T / G) and the reduction rate of the virus was tested, a significant correlation was found with P = 7.2 × 10 ^-17 . (Linear regression model)
from this result
(a) rs8099917 (T / G) genotype in the order of GG>GT> TT, an early study 2 weeks after the start of treatment with IFN therapy (Peginterferon / Ribavirin combination therapy). Low sensitivity to treatment, or
(b) rs8099917 (T / G) genotypes in the order of TT>GT> GG, an early study 2 weeks after the start of treatment with IFN therapy (Peginterferon / Ribavirin combination therapy). It was found that the sensitivity to the treatment could be predicted.

FIG. 4 shows the reduction rate of blood HCV-RNA 4 weeks after the start of IFN therapy (pegylated interferon / ribavirin combination therapy), divided into rs8099917 (T / G) genotypes. The number of patients analyzed was 595.
The horizontal axis is “blood HCV-RNA level 4 weeks after starting IFN therapy (peginterferon / ribavirin combination therapy)” divided by “blood HCV-RNA level before IFN therapy (peginterferon / ribavirin combination therapy)”. The logarithmic conversion value with the base value of 10 as the base value represents the rate of decrease in blood HCV-RNA 4 weeks after the start of IFN therapy (Peginterferon-ribavirin combination therapy). The vertical axis shows the rs8099917 (T / G) genotype in order from the top: (a) GG homozygous type (GG), (b) GT heterozygous type (GT), and (c) TT homozygous type (TT ).
When the correlation coefficient between the genotype of rs8099917 (T / G) and the virus reduction rate was tested, a significant correlation with P = 3.1 X 10 ^-27 was observed (linear regression model).
from this result
(a) rs8099917 (T / G) genotypes in the order of GG>GT> TT, an early study 4 weeks after the start of treatment with IFN therapy (Peginterferon / Ribavirin combination therapy). Low sensitivity to treatment, or
(b) rs8099917 (T / G) genotype in the order of TT>GT> GG, an early study 4 weeks after the start of treatment with IFN therapy (peginterferon / ribavirin combination therapy). It was found that the sensitivity to treatment could be expected.

[Gene polymorphism in linkage disequilibrium with rs8099917]
In order to search for genetic polymorphisms that are in linkage disequilibrium with rs8099917 (T / G), the region of the linkage disequilibrium block where rs8099917 is located was resequenced using genomic DNA of 48 healthy Japanese individuals. Sing was performed.
Among the identified gene polymorphisms, the gene polymorphisms that have a linkage disequilibrium relationship with rs8099917 (T / G) are shown in Table 7 together with r ² indicating a measure of linkage disequilibrium. These results indicate that the gene polymorphisms shown in Table 7 are useful as markers of the present invention and can be used for predicting the therapeutic effect of IFN therapy.

  According to the present invention, it is possible to provide a marker for predicting the therapeutic effect of a patient in IFN therapy, specifically, type I IFN therapy.

[SEQ ID NO: 17]
N represents TT or G.
[SEQ ID NO: 21]
N represents no nucleotide or CT.
[SEQ ID NO: 25]
N represents no nucleotide or GA.
[SEQ ID NO: 26]
N represents AC or no nucleotide.

Claims (17)

The gene polymorphisms present in the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) gene regions located in human chromosome 19 short arm 19q13.13, the following groups:
A) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 1;
B) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 2 (T>A);
C) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 3 (A>T);
D) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 4 (A>G);
E) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 5 (G>A);
F) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 6 (T>C);
G) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 7 (T>C);
H) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 8 (C>T);
I) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 9 (C>A);
J) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 10 (A>G);
K) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 11 (C>G);
L) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 12;
M) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 13 (A>G);
N) a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 14 (C>T);
O) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 15 (A>G);
P) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 16 (C>G);
Q) Gene polymorphism at the 301st base in the nucleotide sequence represented by SEQ ID NO: 17 (TT>G);
R) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 18 (G>C);
S) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 19 (C>A);
T) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 20 (C>T);
U) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 21 (->CT);
V) a gene polymorphism (C> T) at the 301st base in the nucleotide sequence represented by SEQ ID NO: 22;
W) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 23 (A>T);
X) a gene polymorphism (G> A) at the 301st base in the base sequence represented by SEQ ID NO: 24;
Y) gene polymorphism (-> GA) at the 301st base in the base sequence represented by SEQ ID NO: 25; and
Z) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 26 (AC>-);
(However, the parentheses indicate major allele> minor allele, and “-” indicates a base deletion.)
A reagent for predicting susceptibility to treatment with type I interferon, comprising a polynucleotide capable of detecting each minor allele in at least one gene polymorphism selected from:   The reagent according to claim 1, further comprising a polynucleotide capable of detecting each major allele.   A polynucleotide capable of detecting each allele comprises a string of IFNλ2 or IFNλ3 consisting of 10 to 200 consecutive base sequences including the allele in the base sequence represented by each SEQ ID NO. The reagent according to claim 1 or 2, which is a probe capable of hybridizing under mild conditions and / or a primer capable of amplifying the gene fragment.   The reagent according to claim 3, wherein the gene fragment of IFNλ2 or IFNλ3 has a continuous base sequence of 10 to 50.   The reagent according to claim 3, wherein the gene fragment of IFNλ2 or IFNλ3 is composed of 15 to 30 consecutive base sequences.   The reagent according to any one of claims 1 to 5, which is used for predicting whether interferon therapy is effective for a patient having a disease to which treatment with type I interferon can be applied.   The reagent according to claim 6, wherein the patient with a disease to which treatment with type I interferon can be applied is an HCV-infected patient.   The reagent according to claim 7, wherein the HCV infected patient is an HCV genotype 1b infected patient, an HCV genotype 1a infected patient or an HCV genotype 2a infected patient.   The reagent of any one of Claims 1-8 whose HCV infection patient is East Asian. (1) A genetic polymorphism present in the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) gene regions located in human chromosome 19 short arm 19q13.13, using a gene sample derived from a subject, group:
A) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 1;
B) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 2 (T>A);
C) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 3 (A>T);
D) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 4 (A>G);
E) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 5 (G>A);
F) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 6 (T>C);
G) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 7 (T>C);
H) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 8 (C>T);
I) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 9 (C>A);
J) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 10 (A>G);
K) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 11 (C>G);
L) gene polymorphism (C> T) at the 301st base in the base sequence represented by SEQ ID NO: 12;
M) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 13 (A>G);
N) a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 14 (C>T);
O) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 15 (A>G);
P) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 16 (C>G);
Q) Gene polymorphism at the 301st base in the nucleotide sequence represented by SEQ ID NO: 17 (TT>G);
R) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 18 (G>C);
S) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 19 (C>A);
T) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 20 (C>T);
U) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 21 (->CT);
V) a gene polymorphism (C> T) at the 301st base in the nucleotide sequence represented by SEQ ID NO: 22;
W) Gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 23 (A>T);
X) a gene polymorphism (G> A) at the 301st base in the base sequence represented by SEQ ID NO: 24;
Y) gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 25 (->GA);
Z) a gene polymorphism at the 301st base in the base sequence represented by SEQ ID NO: 26 (AC>-); and
AA) gene polymorphism (T> G) at the 301st base in the base sequence represented by SEQ ID NO: 27;
(However, the parentheses indicate major allele> minor allele, and “-” indicates a base deletion.)
And (2) if the presence of a minor allele is found as a result of the test of (2) and (1), the subject is less sensitive to treatment with type I interferon A method for predicting sensitivity of a subject to treatment with type I interferon, comprising a step of determining.   The prediction method according to claim 10, wherein the gene sample includes genomic DNA or RNA.   The prediction method according to claim 10 or 11, wherein the subject is a patient with a disease to which treatment with type I interferon can be applied.   The prediction method according to claim 12, wherein the patient having a disease to which treatment with type I interferon can be applied is an HCV-infected patient.   The prediction method according to claim 13, wherein the HCV-infected patient is an HCV genotype 1b-infected patient, an HCV genotype 1a-infected patient, or an HCV genotype 2a-infected patient.   The prediction method according to any one of claims 10 to 14, wherein the subject is an East Asian. (1) A gene polymorphism present in the interferon λ2 (IFNλ2) and interferon λ3 (IFNλ3) gene regions located in human chromosome 19 short arm 19q13.13, using a gene sample derived from a subject, comprising SEQ ID NO: The step of testing the gene polymorphism at the 301st base in the nucleotide sequence represented by 27, and the results of (2) and (1), the presence of a homozygous gene polymorphism (TT) of a major allele was recognized. A method for predicting the sensitivity of a subject to treatment with type I interferon, comprising the step of determining that the subject is highly sensitive to treatment with type I interferon.   The prediction method according to claim 16, wherein the subject is an East Asian.