T cell proliferation-promoting peptides

Abstract

To provide an inexpensive T-cell growth factor.SOLUTION: A peptide comprises an amino acid sequence selected from the group consisting of specific amino acid sequences, which comprises P, RP, or ERP at N terminus and / or comprises T, TG, TGQ, TGQK, or TGQKP at C terminus.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to a T cell proliferation-promoting peptide. [Background technology]

[0002] In recent years, progress has been made in the development of cancer treatments that utilize a patient's T cells. For example, Kymriah has been approved as a treatment for B-cell acute lymphoblastic leukemia and diffuse large B-cell lymphoma. Kymriah uses genetically modified cells called CAR-T cells, in which a chimeric antigen receptor (CAR) is expressed in healthy T cells collected from the patient. When administered to the patient, these cells selectively attack and eliminate antigen-expressing cancer cells. In this CAR-T cell therapy, the CAR gene is introduced into T cells, and then the cells must be proliferated.

[0003] In order to grow T cells and CAR-T cells in a test tube, it was necessary to add CD3-binding antibodies, CD28-binding antibodies, cytokine IL-2, etc. to the culture medium (Non-Patent Document 1). Because these proteins are all 114 amino acids or longer, they could not be synthesized inexpensively by chemical synthesis, and expensive recombinant proteins had to be used. [Prior art documents] [Non-patent literature]

[0004] [Non-Patent Document 1] Molecular Therapy, 2020, (USA), Vol. 28, pp. 2379-2393 Summary of the Invention [Problem to be solved by the invention]

[0005] If inexpensive T cell growth factors were available for the in vitro proliferation of T cells and CAR-T cells, it would be possible to reduce the manufacturing costs of CAR-T cells and the costs of experiments using T cells in basic research. Therefore, an object of the present invention is to provide an inexpensive T cell growth factor. [Means for solving the problem]

[0006] The present inventors have conducted extensive research into inexpensive T cell growth factors and have surprisingly found that a specific short peptide of about 30 amino acids has potent T cell proliferation activity. The present invention is based on this finding. Therefore, the present invention provides [1] A peptide containing any amino acid at positions 15 to 17 in the amino acid sequence represented by SEQ ID NO: 1, or a peptide containing any amino acid at positions 13 and 20 in the amino acid sequence represented by SEQ ID NO: 2; [2] The peptide according to [1], wherein the amino acid sequence represented by SEQ ID NO: 1 or SEQ ID NO: 2 contains P, RP, or ERP at the N-terminus and / or T, TG, TGQ, TGQK, or TGQKP at the C-terminus. [3] A peptide library comprising the peptide according to [1] or [2]. [4] A peptide comprising an amino acid sequence selected from the group consisting of amino acid sequences represented by SEQ ID NOs: 3 to 62. [5] The peptide according to [4], wherein the amino acid sequence is selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 3 to 62 and contains P, RP, or ERP at the N-terminus and / or T, TG, TGQ, TGQK, or TGQKP at the C-terminus. [6] The peptide according to [4] or [5], wherein the amino acid sequence is the amino acid sequence represented by SEQ ID NO: 5, 6, or 33; and [7] The peptide according to any one of [4] to [6], which is used for T cell proliferation. Regarding. [Effects of the Invention]

[0007] The peptides of the present invention can efficiently promote the proliferation of T cells. The peptides of the present invention have an amino acid chain length of about 30 amino acids and can be chemically synthesized, so they can be produced at low cost. [Brief explanation of the drawings]

[0008] [Figure 1] 1 shows the amino acid sequences of peptides enriched in peptide library 1 by binding to CD25. [Figure 2] 1 shows the amino acid sequences of peptides enriched in peptide library 2 by binding to CD25. [Figure 3] 1 is a graph showing that the addition of peptides 1 to 3 of the present invention promoted the proliferation of human peripheral blood mononuclear cells, as measured by MTT assay. DETAILED DESCRIPTION OF THE INVENTION

[0009] [1] Peptide library The peptide library of the present invention comprises peptides containing any amino acids at positions 15 to 17 in the amino acid sequence represented by SEQ ID NO: 1, or peptides containing any amino acids at positions 13 and 20 in the amino acid sequence represented by SEQ ID NO: 2. The peptides may contain P, RP, or ERP at the N-terminus and / or T, TG, TGQ, TGQK, or TGQKP at the C-terminus of the amino acid sequence represented by SEQ ID NO: 1 or 2.

[0010] The peptides used in the peptide library of the present invention (hereinafter sometimes referred to as "library peptides") are peptides in which a partial sequence of CDR-H3 of a monoclonal antibody against CD25 is incorporated into a scaffold protein. CD25 is one of the three subunits that constitute the IL-2 receptor and is called the α subunit. The inventors selected the α subunit (CD25) from the three subunits of the IL-2 receptor, and then selected CDR-H3 from the six complementarity determining regions of the monoclonal antibody against CD25, namely CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3, and incorporated a partial sequence of CDR-H3 into the scaffold protein.

[0011] The scaffold protein is the one described in Japanese Patent Application No. 2020-206671. YACX1X2X3X4CX5X6X7FX8X9X 10 X 11 X 12 X 13 X 14 A scaffold protein containing RHIRIH (SEQ ID NO: 63) was used. A partial protein of CDR-H3 was incorporated into this scaffold protein. The library peptides prepared were: (1)YACPVESC GGVFDY XXXLTRHIRIH (SEQ ID NO: 1), and (2) YACPVESCDRRFX GGGV LTXHIRIH (SEQ ID NO: 2). The amino acid sequence of the CDR-H3 used was "GGGVFDY (SEQ ID NO: 64)". The library peptide (1) is a peptide in which the partial peptide of CDR-H3, "GGVFDY (SEQ ID NO: 65)", is incorporated into the "X5X6X7FX8X9" region of the scaffold protein. 10 X 11 X 12 " is a peptide containing any amino acid in the region of On the other hand, the library peptide (2) is a scaffold protein "X9X 10 X 11 X 12 " is a peptide incorporating a partial peptide of CDR-H3, "GGGV (SEQ ID NO: 66)," into the region of ". The peptide also contains any amino acid at the positions of "X8" and "R" of the scaffold protein.

[0012] The library peptides of the present invention may contain P, RP, or ERP at the N-terminus of library peptide (1) or library peptide (2) and / or T, TG, TGQ, TGQK, or TGQKP at the C-terminus. Thus, there are four types of N-terminus, including those in which no amino acids are added, and six types of C-terminus, including those in which no amino acids are added. Thus, there are 24 different combinations of N- and C-terminus of library peptide (1) or library peptide (2). Any of these combinations can be used as the peptides in the library of the present invention. Specifically, the 25-amino acid-long library peptide (1) or library peptide (2) can stably present a partial peptide of CDR-H3. Furthermore, 24 types of library peptides in which the amino acids are added to the N-terminus and / or C-terminus and are 26 to 33 amino acids long can also stably present a partial peptide of CDR-H3. For example, library peptides in which "ERP" is added to the N-terminus of library peptide (1) or library peptide (2) and "TGQKP" is added to the C-terminus are each 33 amino acid long. (3)ERPYACPVESC GGVFDY XXXLTRHIRIHTGQKP (SEQ ID NO: 67), and (4)ERPYACPVESCDRRFX GGGV LTXHIRIHTGQKP (sequence number 68).

[0013] The 25-amino acid library peptide (1) or (2) can stably present a partial peptide of CDR-H3 and any amino acid "X." Because the 25-amino acid library peptide (1) or (2) is stable, library peptides of 26 to 33 amino acids in length, including library peptide (1) or (2), can also stably present a partial peptide of CDR-H3 and any amino acid "X." Furthermore, library peptides in which one or more amino acids have been added to the N-terminus and / or C-terminus of the 33-amino acid library peptide (3) or library peptide (4) can also be effectively used as library peptides of the present invention. The number of added amino acids is not particularly limited, but is preferably 1 to 15 (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15), more preferably 1 to 10, even more preferably 1 to 5, and even more preferably 1 to 3. The amino acid to be added is not particularly limited, and may be any of the amino acids "X" described below.

[0014] The "X" amino acid contained in the library peptide is not particularly limited, but may be any of the 20 α-amino acids that constitute proteins in vivo: glycine, alanine, valine, leucine, isoleucine, serine, threonine, cysteine, methionine, asparagine, glutamine, proline, phenylalanine, tyrosine, tryptophan, aspartic acid, glutamic acid, lysine, arginine, or histidine. While the α-amino acids that constitute proteins in vivo are generally L-amino acids, the "X" amino acid may also include D-amino acids.

[0015] Furthermore, although not limited thereto, the arbitrary amino acid "X" can be an amino acid other than the 20 amino acids. That is, the amino acid to be incorporated can be selected depending on the purpose of screening. Specific amino acids include 2,3-diaminopropionic acid, α-aminoisobutyric acid, ε-aminohexanoic acid, δ-aminovaleric acid, N-methylglycine or sarcosine, ornithine, citrulline, t-butylalanine, t-butylglycine, N-methylisoleucine, phenylglycine, cyclohexylalanine, norleucine, naphthylalanine, 2-chlorophenylalanine, 3-chlorophenylalanine, 4-chlorophenylalanine, 2-fluorophenylalanine, 3-fluorophenylalanine, 4-fluorophenylalanine, 2-bromophenylalanine, 3-bromophenylalanine, 4-bromophenylalanine, 2-methylphenylalanine, 3-methylphenylalanine, 4-methylphenylalanine, 2-nitrophenylalanine, 3-nitrophenylalanine, 4-nitrophenylalanine, 2-cyanophenylalanine, 3-cyanophenylalanine, 4 -cyanophenylalanine, 2-trifluoromethylphenylalanine, 3-trifluoromethylphenylalanine, 4-trifluoromethylphenylalanine, 4-aminophenylalanine, 4-iodophenylalanine, 4-aminomethylphenylalanine, 2,4-dichlorophenylalanine, 3,4-dichlorophenylalanine, 2,4-difluorophenylalanine, 3,4-difluorophenylalanine, pyrid-2-ylalanine, pyrid-3-ylalanine, pyrid-4-ylalanine, naphth-1-ylalanine, naphth-2-ylalanine, thiazolylalanine, benzothienylalanine, thienylalanine, furylalanine, homophenylalanine, homotyrosine, homotryptophan, pentafluorophenylalanine, styrylalanine, autorylalanine, 3,3-diphenylalanine, 3-amino-5-phenylpentanoic acid, penicillamine, 1,2,3,Examples of amino acids include 4-tetrahydroisoquinoline-3-carboxylic acid, β-2-thienylalanine, methionine sulfoxide, N(w)-nitroarginine, homolysine, phosphonomethylphenylalanine, phosphoserine, phosphothreonine, homoaspartic acid, homoglutamic acid, 1-aminocyclopent-(2 or 3)-ene-4-carboxylic acid; pipecolic acid, azetidine-3-carboxylic acid, 1-aminocyclopentane-3-carboxylic acid; allylglycine, propargylglycine, homoalanine, norvaline, homoleucine, homovaline, homoisoleucine, homoarginine, N-acetyllysine, 2,4-diaminobutyric acid, 2,3-diaminobutyric acid, N-methylvaline, homocysteine, homoserine, hydroxyproline, and homoproline. The amino acids are preferably α-amino acids, because their structure can stabilize the structure of the library peptides of the present invention. However, even if the scaffold peptide of the present invention contains only a few β-amino acids (e.g., 4 or less, 3 or less, 2 or less, or 1), the structure of the scaffold peptide of the present invention is stable.

[0016] The library peptides of the present invention can be used to screen for peptides that exhibit excellent binding ability to the IL-2 receptor on T cells. That is, since the library peptides of the present invention contain a partial peptide of CDR-H3, they are thought to have a certain degree of affinity for the α subunit (CD25) of the IL-2 receptor, and screening allows for the selection of the amino acid "X" that is optimal for binding, which is thought to enable the selection of peptides with excellent binding ability to the IL-2 receptor on T cells.

[0017] The peptide library of the present invention can be constructed using library techniques known in the art, except that the library peptides of the present invention are used. Polynucleotides encoding the library peptides can be used to construct the peptide library. The polynucleotide is a polynucleotide that encodes a library peptide of the present invention (hereinafter, may be referred to as a library peptide polynucleotide). The library peptide polynucleotide may include a nucleotide that encodes a linker polypeptide. Genes to be incorporated into the library can be prepared by known methods. For example, the NNK codon (where N is any of the bases A, G, C, or T, and K is any of the bases G or T) encoding any amino acid in "X" can be site-specifically replaced with a codon encoding any amino acid. Substitution can be achieved by chemically synthesizing an oligo DNA containing the desired region and incorporating it using a restriction enzyme, or by replicating the entire vector using PCR.

[0018] The polynucleotides can be easily amplified by, but not limited to, incorporating them into a vector, and can be easily expressed as library peptides. The vector into which the polynucleotide is inserted can be any vector, such as a plasmid, phage, or virus, as long as it is replicable in host cells. Examples include Escherichia coli plasmids such as pBR322, pBR325, pUC118, pUC119, pKC30, and pCFM536; Bacillus subtilis plasmids such as pUB110; yeast plasmids such as pG-1, YEp13, and YCp50; and phage DNA such as λgt110 and λZAPII. Vectors for mammalian cells include viral DNA such as baculovirus, vaccinia virus, adenovirus, and lentivirus, as well as SV40 and its derivatives. The vector contains a replication origin, a selection marker, and a promoter, and may also contain an enhancer, a transcription termination sequence (terminator), a ribosome binding site, a polyadenylation signal, and the like, as necessary.

[0019] As used herein, the term "host" is not limited as long as the vector can be integrated and the library peptide can be expressed. For example, hosts for phage vectors include M13 phage, fd phage, and T7 phage. Examples of bacterial vector hosts include Escherichia coli, Streptomyces, and Bacillus subtilis. Examples of yeast vector hosts include baker's yeast and methylotrophic yeast. Examples of insect cell vector hosts include Drosophila S2 and Spodoptera Sf9. Examples of animal cell vector hosts include CHO, COS, BHK, 3T3, and C127.

[0020] Phage surface proteins include, but are not limited to, g3p and g8p of M13 phage, or G10 of T7 phage. Bacterial surface proteins, for example, E. coli surface proteins, include, but are not limited to, OmpA. Yeast surface proteins include, but are not limited to, Flo1, alpha agglutinin, or AGA2. Examples of animal cell surface proteins include, but are not limited to, human Toll-like receptor 4, EGFR, LDL receptor, angiotensin II receptor, or PDGFR.

[0021] Therefore, when affinity selection (screening) of a target peptide is performed using the peptide library of the present invention, the target peptide can be easily concentrated. The screening method comprises (1) the step of contacting CD25 with a peptide library, and (2) the step of recovering hosts that bind to CD25, and preferably further comprises (3) the step of amplifying the recovered hosts. The screening method preferably repeats the contacting step (1), the host recovery step (2), and the amplification step (3) to enrich peptides that bind to CD25. The contacting of CD25 with the peptide library in the contacting step (1) can be carried out by immobilizing the target molecule on an insoluble carrier (e.g., a well or a dish). The screening method can be performed according to known methods, except that the peptide library of the present invention is used. For example, screening of a phage library using phages as hosts can be performed as follows: A target molecule immobilized on a plate or the like is contacted with the phage library to allow a reaction. Phage that do not bind to the target molecule are removed by washing. Phage that bind to the target molecule are recovered, infected with E. coli, lysed, and allowed to grow. The culture supernatant after lysis or purified phages are further reacted with immobilized target molecules (panning) to concentrate phages that display binding molecules specific to the target molecule. Finally, the phages are cloned, and the displayed peptides are identified by sequencing. The obtained peptides are peptides that can bind to CD25. In the screening method, since the peptides are presented on the scaffold protein, the peptide structure is stable, making it possible to screen for peptides that are highly specific to CD25.

[0022] [2] Peptides for T cell proliferation The peptides of the present invention are peptides (hereinafter sometimes referred to as T cell proliferation peptides) comprising an amino acid sequence selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 3 to 62, and exhibit high affinity for CD25 and have the activity of proliferating T cells. The T cell proliferation sequences of the amino acid sequences represented by SEQ ID NOs: 3 to 62 are shown in Tables 1 and 2.

[0023] [Table 1]

[0024] [Table 2]

[0025] The T cell proliferation peptides of the present invention may contain P, RP, or ERP at the N-terminus and / or T, TG, TGQ, TGQK, or TGQKP at the C-terminus in the amino acid sequences represented by SEQ ID NOS: 3 to 62. Thus, there are four types of N-terminus, including those in which no amino acids are added, and six types of C-terminus, including those in which no amino acids are added. Therefore, there are 24 different combinations of N- and C-terminus of T cell proliferation peptides. Any of these combinations can be used as the T cell proliferation peptide of the present invention. Specifically, the 25-amino acid T cell proliferation peptides stably exhibit high affinity for CD25 and exhibit T cell proliferation activity. Furthermore, 24 types of library peptides 26 to 33 amino acids in length, in which the amino acids are added to the N-terminus and / or C-terminus, also stably exhibit high affinity for CD25 and exhibit T cell proliferation activity.

[0026] For example, a T cell proliferation peptide having "ERP" added to the N-terminus and "TGQKP" added to the C-terminus of the T cell proliferation peptide is obtained, each having a length of 33 amino acids. Peptide 1: ERPYACPVESCGGVFDYHIGLTRHIRIHTGQKP (SEQ ID NO: 69), Peptide 2: ERPYACPVESCGGVFDYKVQLTRHIRIHTGQKP (SEQ ID NO: 70), or Peptide 3: ERPYACPVESCDRRFTGGGVLTAHIRIHTGQKP (SEQ ID NO: 71).

[0027] The 25-amino acid T cell proliferation peptides stably exhibit high affinity for CD25 and exert T cell proliferation effects. Because these 25-amino acid T cell proliferation peptides are stable, 26- to 33-amino acid T cell proliferation peptides, including these T cell proliferation peptides, also stably exhibit high affinity for CD25 and exert T cell proliferation effects. Furthermore, library peptides in which one or more amino acids have been added to the N-terminus and / or C-terminus of the 33-amino acid T cell proliferation peptides can also be effectively used as library peptides of the present invention. The number of added amino acids is not particularly limited, but is preferably 1 to 15 (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15), more preferably 1 to 10, even more preferably 1 to 5, and even more preferably 1 to 3.

[0028] The equilibrium dissociation constant (K D ) is not particularly limited, but is, for example, 0.001 to 10,000K D (nM), and in one embodiment, 0.01 to 100K D (nM), and in one embodiment, 1 to 100K D (nM), and in one embodiment, 10 to 80K D (nM), and in one embodiment, 20 to 60K D (nM). When the equilibrium dissociation constant is in the above range, the antibody can efficiently bind to CD25 and proliferate T cells.

[0029] 《Action》 The T cell proliferation peptides of the present invention have a small molecular size and do not contain intramolecular disulfide bonds, and therefore can be easily prepared by chemical synthesis. [Example]

[0030] The present invention will be specifically described below with reference to examples, but these examples are not intended to limit the scope of the present invention.

[0031] Example 1 In this example, a peptide library having the following two types of amino acid sequences was constructed, in which partial peptides of CDR-H3 of a monoclonal antibody against CD25 were incorporated into a scaffold peptide. Peptide Library 1: ERPYACPVESC GGVFDYXXX LTRHIRIHTGQKP (SEQ ID NO: 67) Peptide Library 2: ERPYACPVESCDRRF XGGGVLTX HIRIHTGQKP (SEQ ID NO: 68) For peptide library 1, amino acids 12 to 20 of the scaffold peptide (ERPYACPVESCDRRFSRSDELTRHIRIHTGQKP: SEQ ID NO: 72) were replaced with "GGVFDYXXX" for "DRRFSRSDE." For peptide library 2, amino acids 16 to 23 of "SRSDELTR" were replaced with "XGGGVLTX." Using a vector containing nucleic acid encoding the scaffold peptide as a template, nucleotides encoding "ERPYACPVESCGGVFDYXXXLTRHIRIHTGQKP" or "ERPYACPVESCDRRFXGGGVLTXHIRIHTGQKP" were synthesized by PCR. Nucleotides (primers) were synthesized so that the X amino acid was an NNK codon (where N is A, G, C, or T, and K is G or T), which encodes any of the 20 amino acids, and used in the reaction. The resulting nucleotides were excised with restriction enzymes EcoRI and XhoI and inserted into a lentiviral vector to obtain peptide libraries 1 and 2.

[0032] Example 2 In this example, peptide libraries 1 and 2 constructed in Example 1 were used to perform screening (enrichment) based on binding to CD25. First, a K562 cell line that constitutively expresses CD25 was established, and the library peptides obtained in Example 1 were co-expressed on the cell membrane. From this cell line, a group of cells in which CD25 bound to the peptides was collected, the genome was extracted, and the gene sequences encoding the library peptides were analyzed using a next-generation sequencer. The top 30 peptide sequences from peptide library 1 are shown in Figure 1, and the top 30 peptide sequences from peptide library 2 are shown in Figure 2. These peptides were enriched by binding to CD25 and exhibit excellent affinity for CD25.

[0033] Example 3 In this example, the peptides obtained in Example 2 were chemically synthesized, and their binding strength to CD25 was measured by biolayer interferometry. The following peptides 1 to 3 were chemically synthesized. Peptide 1: ERPYACPVESCGGVFDYHIGLTRHIRIHTGQKP (SEQ ID NO: 69) Peptide 2: ERPYACPVESCGGVFDYKVQLTRHIRIHTGQKP (SEQ ID NO: 70) Peptide 3: ERPYACPVESCDRRFTGGGVLTAHIRIHTGQKP (SEQ ID NO: 71) The peptides were modified with biotin at their N-terminus and diluted to 100 nM in buffer (TBS buffer containing 0.1% BSA and 0.002% Tween-20, pH 7.4). The peptides were loaded onto the surface of a streptavidin biosensor for 300 seconds. After washing the biosensor for 30 seconds and equilibrating for 60 seconds, binding of CD25-Fc to the peptides immobilized on the biosensor was measured for 300 seconds. The biosensor was then immersed in buffer, and dissociation of CD25-Fc was measured for 300 seconds. The resulting binding curves were analyzed to determine the equilibrium dissociation constant (KD). As shown in Table 3, peptides 1 to 3 exhibited excellent equilibrium dissociation constants.

[0034] [Table 3]

[0035] Example 4 In this example, the proliferation activity of human T cells was examined using peptides 1 to 3. Peripheral mononuclear cells were isolated from blood collected from volunteers using Lymphoprep. 5Human peripheral blood mononuclear cells were cultured, and the peptides 1 to 3 (10 μg / mL) were added to the culture medium. IL-2 (200 U / mL) was added as a positive control, and the scaffold peptide (10 μg / mL: the amino acid sequence is shown below) was added as a negative control. Scaffold peptide: ERPYACPVESCDRRFSRSDELTRHIRIHTGQKP (SEQ ID NO: 72) After 7 days of culture, the cell number was measured by MTT assay (n=3). As shown in Figure 3, the addition of peptides 1, 2, and 3 significantly increased the cell number. This cell proliferation effect was stronger than that of IL-2. [Industrial Applicability]

[0036] The T cell proliferation peptide of the present invention has the effect of proliferating T cells and can be used, for example, to proliferate T cells in CAR-T cell therapy.

Claims

1. A peptide library for screening peptides for T cell proliferation, comprising peptides containing any amino acid at positions 15 to 17 in the amino acid sequence represented by SEQ ID NO: 1, or peptides containing any amino acid at positions 13 and 20 in the amino acid sequence represented by SEQ ID NO:

2.

2. The peptide library described in claim 1, wherein the peptides containing any amino acid at positions 15 to 17 in the amino acid sequence represented by SEQ ID NO: 1, or any amino acid at positions 13 and 20 in the amino acid sequence represented by SEQ ID NO: 2, are peptides that contain P, RP, or ERP at the N-terminus and / or T, TG, TGQ, TGQK, or TGQKP at the C-terminus.

3. A peptide comprising an amino acid sequence selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 5, 6, and 33.

4. The peptide of claim 3, which has an amino acid sequence selected from the group consisting of the amino acid sequences represented by SEQ ID NOs: 5, 6, and 33, and which contains P, RP, or ERP at the N-terminus and / or T, TG, TGQ, TGQK, or TGQKP at the C-terminus.

5. The peptide according to claim 3 or 4, which is used for T cell proliferation.

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