JP2019080491A - Method for inducing NY-ESO1 antigen-specific T cells for immune cell therapy - Google Patents

Abstract

To provide methods for inducing T cells for immune cell therapy in an immune cell therapy for allotransplanting NY-ESO1 antigen specific T cells.SOLUTION: The present invention provides a method for inducing T cells for immune cell therapy comprising (1) providing human pluripotent stem cells having NY-ESO1 antigen-specific T cell receptor, and (2) inducing pre-T cells or mature T cells from the pluripotent stem cells of the step (1). The human pluripotent stem cells having NY-ESO1 antigen-specific T cell receptor is preferably prepared by regenerating T cells from T-iPS cells, which are iPS cells induced from NY-ESO1 antigen-specific T cells isolated or induced from a human.SELECTED DRAWING: None

Description

  The present application relates to a method of inducing T cells for immunocell therapy. In particular, the present application relates to a method for inducing T cells for immune cell therapy in immune cell therapy for allotransplanting NY-ESO1 antigen-specific T cells.

Individual T cells express T cell receptors (TCRs) of different specificities. When an infection occurs, cells of a specific specificity proliferate and form cell populations (clones) to address pathogens. This is a basic form of the acquired immune system. It is expected that T cells of a specific specificity can be artificially expanded (referred to as cloning) and used for therapy. In fact, T cells showing specific specificity are collected from patients, expanded and returned to patients (autologous transplantation). However, most of such attempts have not been purified as much as cloning, and there has also been a problem that the activity to kill cancer cells decreases while being subcultured for several generations in vitro .

  A method of infinitely expanding T cells by immortalizing them has also been proposed. One cell is immortalized, expanded and cloned. Immortalization of cells includes a method by fusion with cancer cells and a method such as long-term culture by TCR stimulation and cytokine stimulation. However, such immortalized T cells are, so to speak, cancer cells, and autologous transplantation for returning to the patient itself is dangerous. In addition, there is also a problem that the function is deteriorated in the cloning process.

  NY-ESO1 is a cancer / testis antigen which expression is observed in melanoma, esophageal cancer, multiple myeloma, and part of adult T cell leukemia (ATL) but not in normal tissues except in testis. The NY-ESO1 antigen is one of the most clinically focused and studied antigens as cancer antigens. Clinical trials have been conducted for cancer vaccine therapy using NY-ESO1 antigen peptide.

The NY-ESO1 antigen-specific T cell receptor (TCR = T cell receptor) gene has been isolated. The clinical trials have been carried out for gene therapy that expresses the TCR gene in the patient's normal T cells (aggregate of many clones) and returns it back to the patient's body (autologous transplantation), and is effective in specific cancer types It has been reported. (Non-patent documents 1 to 3). At the time of expressing a TCR, suppression of the TCR originally expressed by a patient's normal T cell is carried out by siRNA or the like.
Ru.

  There are at least three problems with this TCR gene therapy: 1) Transfer of TCR gene is carried out using a retrovirus vector, that is, it is gene therapy, and it is possible for T cells of the transferred patient to become cancerous. 2) The suppression of endogenous TCR by siRNA is not complete, and unexpected reactivity that attacks the tissues of the patient by swapping etc. between various endogenous TCRa chains or TCRb chains and the introduced TCRa chain or TCRb chain There is a risk of 3) It is necessary to recover T cells, genetically modify, and administer for each patient, it is not possible to prepare in advance, and treatment takes time.

WO2011 / 096482 WO 2016/010153 WO 2016/010154 WO 2016/010155

Robbins PF et al., J Clin Oncol. 2011; 29 (7): 917-924 Robbins PF et al., Clin Cancer Res. 2015; 21 (5): 1019-1027 Chapuis et al, Sci Transl Med, 5: 174ra27, 2013 Vizcardo et al., Cell Stem Cell. 12: 31-36. 2013. Takahashi and Yamanaka, Cell 126, 663-673 (2006) Takahashi et al., Cell 131, 861-872 (2007) Grskovic et al., Nat. Rev. Drug Dscov. 10, 915-929 (2011) Anticancer Research 32 (12); 5201-5209, 2012 Jurgens et al, Journal of Clinical Investigation, 26:22, 2006 Morgan R. A. et al, Science, 314: 126. 2006 Timmermans et al., Journal of Immunology, 2009, 182: 6879-6888 Blood 111: 1318 (2008) Nature Immunology 11: 585 (2010)

  An object of the present application is to provide a method for inducing T cells for immune cell therapy in immune cell therapy for transplantation of NY-ESO1 antigen-specific T cells into allogeneic transplantation. The present application further aims to provide an immune cell therapy using T cells derived from the pluripotent stem cells.

  The present application also aims to provide pluripotent stem cells having a T cell receptor gene specific for NY-ESO1 antigen.

The present application provides (1) providing human pluripotent stem cells having a T cell receptor gene specific to NY-ESO1 antigen, and (2) inducing T cells from pluripotent stem cells in step (1). Provided a method of inducing T cells for immunocell therapy.

  In a first aspect of the present application, pluripotent stem cells having a T cell receptor gene specific for NY-ESO1 antigen isolate or induce NY-ESO1 antigen specific T cells from a donor, from said T cells Induce pluripotent stem cells. When iPS cells are induced as pluripotent stem cells, iPS cells derived from T cells are referred to as T-iPS cells. The donor of NY-ESO1 antigen-specific T cells may be a healthy person, a healthy person having the NY-ESO1 antigen, or a cancer patient.

  In a second aspect of the present application, pluripotent stem cells having a T cell receptor gene specific to NY-ESO1 antigen introduce T cell receptor gene specific to NY-ESO1 antigen into pluripotent stem cells. Can be obtained by

  The pluripotent stem cells having a T cell receptor gene specific to the obtained NY-ESO1 antigen, T precursor cells or T cells which are mature T cells are induced from T-iPS cells, and used for immune cell therapy .

  T precursor cells or mature T cells obtained by the method of the present application can be used for cell immunotherapy. In cellular immunotherapy, the mature T cells of the present invention can be used particularly preferably not only in autologous transplantation, but also in treatment by allogeneic transplantation to other persons who have a certain HLA match or more.

  The idea of transplanting T cells from other people is contrary to the conventional wisdom of immune cell therapy. For example, in hematological malignancies (such as leukemia), bone marrow transplantation for transplanting hematopoietic stem cells is carried out, but usually from an HLA type donor consistent with the recipient such that the donor bone marrow is not rejected by the recipient. Ru. However, in other humans, the amino acid sequences of many protein molecules other than HLA are mismatched, and donor T cells can recognize these mismatches as an attack target. As a result, a so-called graft-versus-host reaction occurs, which is a reaction in which some of the transplanted donor T cells attack the cells of the recipient's body. If the HLAs do not match, this graft versus host reaction occurs more strongly. These are in fact frequently found after bone marrow transplantation. From such a background, it is common knowledge among medical personnel that "it is dangerous to transplant another person's T cells".

  However, problems with conventional art recognition can be unexpectedly solved by using T cells derived from pluripotent stem cells having a specific antigen-specific T cell receptor gene obtained by the present invention. .

  That is, since the T-iPS cell method which is the first aspect of the present application is not gene therapy, there is no risk of cell carcinogenesis caused by gene transfer. In addition, in CTLs regenerated from iPS cells, endogenous TCR rearrangement is suppressed and only the introduced TCR is expressed, so there is little risk of an unexpected risk. In addition, it is safer to control the expression of endogenous TCR genes at the iPS stage. Since TCR-iPS cells can be treated as clones, identifying a gene insertion site and determining and using a safe clone can avoid the risk of damaging the gene.

Furthermore, in the second aspect of the present application, the following effects can be obtained since it is obtained by the method including the step of introducing the NY-ESO1 antigen-specific T cell receptor gene (TCR gene) into pluripotent stem cells:
1) By introducing the gene of TCR whose effect and safety are guaranteed, the quality of the obtained T cell for transplantation is guaranteed.
2) It is possible to identify the TCR gene insertion site, to determine and use safe clones, and to avoid the problem of canceration of transplanted cells in advance.
3) When differentiating pluripotent stem cells obtained by inserting the TCR gene into pluripotent stem cells into T cells, the cells originally possess since the introduced TCR is first expressed in the differentiation process (hereinafter referred to as endogenous) The TCR is not reconstituted, and unexpected reactions rarely occur.

  In the present application, pluripotent stem cells having a specific antigen-specific T cell receptor are generated, and T precursor cells or mature T cells are derived from the pluripotent stem cells and used for cell immunotherapy. It has been found for the first time that adopting this method does not cause graft vs. host reaction because the cells to be transplanted become T cells with single specificity, and that not only autologous transplantation but also allotransplantation is possible. The That is, only by the present application, it becomes possible to "transplant T cells to another person".

  In the treatment methods for transplanting cells and tissues other than T cells which have been induced to differentiate from various iPS cells, which have been proposed up to now, it is expected that the cells will continue to be engrafted for life. Therefore, when transplanting cells induced to differentiate from iPS cells in the iPS cell stock business premised on allotransplantation, patients need to keep drinking the immunosuppressant. This point is a disadvantage as compared to autologous iPS cells. However, in the case of T-iPS or TCR-iPS cell-derived T cells in the present application, they are rejected after a certain period of time. That is, although the HLA matches more than a certain amount between the donor and the recipient, in other people the minor histocompatibility antigens do not match, and eventually they are rejected. In this respect, unexpected superior effects are shown, which is quite different from other cell transplantation using iPS cells.

  Both T-iPS cells and TCR-iPS cells need not be prepared for each patient, and many types of T-iPS cells and TCR-iPS cells can be prepared in advance. Thus, it is possible to bank at the level of T-iPS cells, TCR-iPS cells, or T precursor cells or T cells derived from such cells. In other words, the construction of a system for enforcing "transplant T cells to another person" for the first time is greatly accelerated by this application. In addition to shortening the time to treatment, there is also the advantage that the quality of transplanted cells can be confirmed before transplantation.

The FACS analysis result which shows that NY-ESO1 tetramer positive and CD8 positive T cells were induced from healthy human volunteers origin T cells in Example 1. A photograph of iPS cell colonies derived from NY-ESO1 antigen peptide-specific T cells. T-iPS cells derived from NY-ESO1 antigen peptide-specific T cells were induced to differentiate into T cells. By FACS analysis of the obtained cells, it was confirmed that the cells were NY-EOS1 tetramer positive and CD3 positive cells. It NY-ESO1 tetramer positive CD3-positive cells in FIG. 3 is a CD8 single positive cells (left) and CD8α ⁺ CD8β ⁺ to have a surface trait (right) was observed. The peptide-specific cytotoxic activity with respect to LCL of the mature T cell (CTL) regenerated from the T-iPS cell derived from NY-ESO1 antigen peptide specific T cell is shown. An adult T-cell leukemia strain in which mature T cells (CTL) regenerated from T-iPS cells derived from NY-ESO1 antigen peptide-specific T cells of A0201-positive healthy individuals express A0201-positive and NY-ESO-1 It confirmed that it had cytotoxic activity with respect to TL-Om1, and that such activity was suppressed by anti-HLA antibody (dotted line). Mature T cells (CTL) regenerated from T-iPS cells derived from T cells specific to NY-ESO1 antigen peptide of A0201-positive healthy individuals are NY-ESO-1-positive adult T cell leukemia strains not expressing A0201 It was confirmed that it did not have cytotoxic activity against ATN-1. Mature T cells (CTL) regenerated from T-iPS cells derived from T cells specific to NY-ESO1 antigen peptide of A0201-positive healthy individuals are NY-ESO-1-positive adult T cell leukemia strains not expressing A0201 It was confirmed that it did not have cytotoxic activity against TL-Su. A multiple myeloma cell line in which mature T cells (CTLs) regenerated from T-iPS cells derived from NY-ESO1 antigen peptide-specific T cells of A0201-positive healthy individuals express A0201-positive and NY-ESO-1 It was confirmed that it had cytotoxic activity against U266, and that such activity was suppressed by anti-HLA antibody (dotted line). NY-ESO-1 positive multiple myeloma cell line in which mature T cells (CTL) regenerated from T-iPS cells derived from NY-ESO1 antigen peptide-specific T cells of A0201 positive healthy individuals do not express A0201 It was confirmed that it did not have cytotoxic activity against RPMI 8226. NY-ESO-1 positive multiple myeloma cell line in which mature T cells (CTL) regenerated from T-iPS cells derived from NY-ESO1 antigen peptide-specific T cells of A0201 positive healthy individuals do not express A0201 It was confirmed that it did not have cytotoxic activity against KMS 28 BM. An adult T-cell leukemia strain in which mature T cells (CTL) regenerated from T-iPS cells derived from NY-ESO1 antigen peptide-specific T cells of A0201-positive healthy individuals express A0201-positive and NY-ESO-1 Survival of mice implanted with TL-Om1 was prolonged. A multiple myeloma cell line in which mature T cells (CTLs) regenerated from T-iPS cells derived from NY-ESO1 antigen peptide-specific T cells of A0201-positive healthy individuals express A0201-positive and NY-ESO-1 Survival of mice implanted with U266 was prolonged.

  In the present specification and claims, “pluripotent stem cells” are stem cells having pluripotency capable of differentiating into many cells present in a living body and having a self-proliferating ability. Pluripotent stem cells include, for example, embryonic stem (ES) cells, cloned embryonic derived embryonic stem (ntES) cells obtained by nuclear transfer, embryonic germ cells ("EG cells"), induced pluripotent stems ( Examples include iPS cells, cultured fibroblasts, and pluripotent cells (Muse cells) derived from bone marrow stem cells. The pluripotent stem cells are preferably mammalian pluripotent stem cells, more preferably human pluripotent stem cells. It is preferable to use iPS cells in consideration of producing a cell bank of a therapeutic method using human-derived cells having a specific HLA.

  In the present specification and claims, "T cell" means a cell expressing on its surface an antigen receptor which is to be a T cell receptor (TCR). It is reported, for example, in WO 2011/096482 and Vizcardo et al., Cell Stem Cell 12, 31-36 2013 that TCR is maintained even when iPS cells are induced from T cells.

In the first aspect of the present application, iPS cells are obtained from desired antigen-specific T cells. The T cells which are induced to iPS cells are preferably T cells which express CD3 and which express at least one molecule selected from the group consisting of CD4 and CD8. As such human T cells, for example, helper / regulatory T cells which are CD4 positive cells, cytotoxic T cells which are CD8 positive cells, naive T cells (CD45RA ⁺ CD62L ⁺ cells), central memory T cells (CD4 positive cells) CD45RA ^- CD62L ⁺ cells), effector memory T cells (CD45RA ^- CD62L ^- cells), and terminal effector T cells (CD45RA ⁺ CD62L ^- cells).

  Human T cells can be isolated from human tissues by known techniques. The human tissue is not particularly limited as long as it is a tissue containing the T cell, and examples thereof include peripheral blood, lymph nodes, bone marrow, thymus, spleen, cord blood, and lesion tissue. Among these, peripheral blood and umbilical cord blood are preferred in view of low invasiveness to humans and easy preparation. Known techniques for isolating human T cells include, for example, flow cytometry using an antibody against a cell surface marker such as CD4 as shown in the following example and a cell sorter. In addition, desired T cells can be isolated using cytokine secretion or expression of functional molecules as an indicator. In such a case, for example, since T cells differ in cytokines secreted by either Th1 type or Th2 type, such cytokines can be used as an index to isolate T cells having a desired Th type. . In addition, cytotoxic (killer) T cells can be isolated by using secretion or production of granzyme or perforin as an indicator.

  The cytotoxic T cells specific for NY-ESO1 antigen can be obtained by stimulating lymphocytes obtained by a conventional method from human, for example, peripheral blood mononuclear cells, with NY-ESO1 antigen or its epitope peptide. A plurality of epitope peptides of the NY-ESO1 antigen have been identified, and methods for inducing NY-ESO1 antigen-specific cytotoxic T cells are also well known. Alternatively, lymphocytes may be stimulated using cancer cells expressing NY-ESO1 antigen.

The NY-ESO1 antigen-specific cytotoxic T cells may be derived from cells of a patient suffering from or expressing a subject expressing a NY-ESO1 antigen, or from a healthy individual. Good. When using T-iPS cells obtained from T cells of healthy individuals, the following effects can be obtained:
1) Since various antigen-specific T cells can be derived from cells of healthy individuals, T-i PS cells having many kinds of TCR genes can be prepared in advance.
2) Targeting healthy individuals, it is easy to collect donors in making T-iPS banks.

  In general, antigen-specific T cells are considered to be collected from patients with infectious diseases and cancer. That is because antigen-specific T cells are amplified in the body of patients with infections and cancer, and it is thought that it is easy to detect / collect specific reactive T cells. The present application provides a method for collecting T cells specific for an antigen associated with a disease from a patient having such a disease and using it as a material for T-iPS cells for allogeneic transplantation.

  In the case of isolating a specific antigen-specific human T cell, a method for purifying the target antigen from a human cultured cell or human tissue containing NY-ESO1 antigen-specific T cell using an affinity column etc. It can be adopted. In addition, it is possible to use a tetramerized MHC (major histocompatibility complex) to which a desired antigen is bound (so-called "MHC tetramer") to use a specific antigen specificity, for example, A method of purifying T cells having specificity for ESO1 antigen can also be employed.

  Pluripotent stem cells are derived from NY-ESO1 antigen-specific T cells obtained as described above. In order to obtain pluripotent stem cells from T cells, for example, the method described in Vizcardo et al., Cell Stem Cell 12, 31-36 2013 may be employed. For example, desired antigen-specific T cells can be obtained from a subject who has acquired immunity to a disease to be treated, and Yamanaka factor can be introduced into these cells to obtain T-iPS cells (Takahashi and Yamanaka, Cell 126, 663) -673 (2006), Takahashi et al., Cell 131, 861-872 (2007) and Grskovic et al., Nat. Rev. Drug Dscov. 10, 915-929 (2011)).

  An induced pluripotent stem (iPS) cell is an artificial somatic stem cell derived from somatic cells, which can produce specific reprogramming factors by acting on somatic cells and has almost the same characteristics as ES cells ( K. Takahashi and S. Yamanaka (2006) Cell, 126: 663-676; K. Takahashi et al. (2007), Cell, 131: 861-872; J. Yu et al. (2007), Science, 318: 1917-1920; Nakagawa, M. et al., Nat. Biotechnol. 26: 101-106 (2008); International Publication WO 2007/069666). The reprogramming factor may be a gene specifically expressed in ES cells, its gene product or non-cording RNA, or a gene that plays an important role in the maintenance of undifferentiated ES cells, its gene product or non-coding RNA, or You may be comprised by the low molecular weight compound. As genes included in the reprogramming factor, for example, Oct3 / 4, Sox2, Sox1, Sox3, Sox15, Sox17, Klf4, Klf2, c-Myc, N-Myc, L-Myc, Nanog, Lin28, Fbx15, ERas, ECAT15 -2, Tcl1, beta-catenin, Lin28b, Sall1, Sall4, Esrrb, Nr5a2, Tbx3 or Glis1 etc. are exemplified, and these reprogramming factors may be used alone or in combination. As a combination of initialization factors, WO2007 / 069666, WO2008 / 118820, WO2009 / 007852, WO2009 / 032194, WO2009 / 058413, WO2009 / 057831, WO2009 / 075119, WO2009 / 079007, WO2009 / 091659, WO2009 / 101084, WO2009 / 01084 101407, WO2009 / 102983, WO2009 / 114949, WO2009 / 126450, WO2009 / 126250, WO2009 / 126251, WO2009 / 126655, WO2009 / 157593, WO2010 / 009015, WO2010 / 033906, WO2010 / 033920, WO2010 / 042800, WO2010 / 050626, WO 2010/056831, WO 2010/068955, WO 2010/098419, WO 2010/102267, WO 2010/111409, WO 2010/111422, WO 2010/115050, WO 2010/124290, WO 2010/147395, WO 2010/147612, Huangfu D, et al. 2008), Nat. Biotechnol., 26: 795-797, Shi Y, et al. (2008), Cell Stem Cell, 2: 525-528, Eminli S, et al. (2008), Stem Cells. 26: 2467. -2474, Huangfu D, et al. (2008), Nat Biotechnol. 26: 1269-1275, Shi Y, et al. (2008), Cell Stem Cell, 3, 568-574, Zhao Y, et al. (2008) ), Cell Stem Cell, 3: 475-479, Marson A, (2008), Cell Stem Cell, 3, 132-135, Feng B, et al. (2009), Nat Cell Biol. 11: 197-203, RL Judson et al., (2009), Nat. Biotechnol., 27: 459-461, Lyssiotis CA, et al. (2009), Proc Natl Acad Sci US A. 106: 8912-8917, Kim JB, et al. (2009), Nature. 461: 649-643, Ichida JK, et al. (2009), Cell Stem Cell. 5: 491-503, Heng J C, et al. (2010), Cell Stem Cell. 74, Han J, et al. (2010), Nature. 463: 1096-100, Mali P, et al. (2010), Stem Cells. 28: 713-720, Maekawa M, et al. (2011), Nature The combinations described in 474: 225-9 are exemplified. (The documents in this paragraph constitute part of the present application by citation)

  The reprogramming factor may be brought into contact with or introduced into somatic cells by a known method depending on its form.

  When in the form of a protein, it may be introduced into somatic cells by techniques such as lipofection, fusion with cell membrane permeable peptides (eg, TAT and polyarginine derived from HIV), microinjection and the like.

  In the case of the form of DNA, for example, it can be introduced into somatic cells by techniques such as viruses, plasmids, vectors such as artificial chromosomes, lipofection, liposomes, microinjection and the like. Examples of viral vectors include retrovirus vectors and lentivirus vectors (above, Cell, 126, pp. 663-676, 2006; Cell, 131, pp. 861-872, 2007; Science, 318, pp. 1917-1920, 2007 And adenovirus vectors (Science, 322, 945-949, 2008), adeno-associated virus vectors, Sendai virus vectors (WO 2010/008054) and the like. Moreover, examples of artificial chromosome vectors include human artificial chromosome (HAC), yeast artificial chromosome (YAC), bacterial artificial chromosome (BAC, PAC) and the like. As a plasmid, a plasmid for mammalian cells can be used (Science, 322: 949-953, 2008). The vector can contain regulatory sequences such as a promoter, enhancer, ribosome binding sequence, terminator, polyadenylation site, etc. so that the nuclear reprogramming substance can be expressed, and further, if necessary, a drug resistance gene ( For example, kanamycin resistance gene, ampicillin resistance gene, puromycin resistance gene, etc.), thymidine kinase gene, selective marker sequences such as diphtheria toxin gene etc., reporter gene sequences such as green fluorescent protein (GFP), β-glucuronidase (GUS), FLAG etc. Can be included. In addition, after the vector is once introduced into the somatic cell and allowed to act, the gene or promoter encoding the reprogramming factor and the gene encoding the reprogramming factor linked thereto are excised together before and after them. It may have a LoxP sequence. (The documents in this paragraph constitute part of the present application by citation)

  When the reprogramming factor is in the form of RNA, it may be introduced into somatic cells by a technique such as lipofection or microinjection. In order to suppress degradation, RNA incorporated with 5-methylcytidine and pseudouridine (TriLink Biotechnologies) may be used as a reprogramming factor (Warren L, (2010) Cell Stem Cell. 7: 618-630). (The documents in this paragraph constitute part of the present application by citation)

  The culture solution for iPS cell induction is, for example, DMEM, DMEM / F12 or DME culture solution containing 10-15% FBS (LIF, penicillin / streptomycin, puromycin, L-glutamine, and the like). Non-essential amino acids, β-mercaptoethanol etc. may be suitably included) or commercially available culture fluid [eg, culture fluid for mouse ES cell culture (TX-WES culture fluid, Thrombox X), primate ES cell culture Medium (culture solution for primate ES / iPS cells, Reprocel), serum-free medium (mTeSR, Stemcell Technology), etc. are exemplified.

As an example of iPS cell induction, for example, a somatic cell is contacted with a reprogramming factor on DMEM or DMEM / F12 culture medium at 10% FBS in the presence of 5% CO ₂ at 37 ° C. for about 4 to 7 days After culture, the cells are replated on feeder cells (eg, mitomycin C-treated STO cells, SNL cells, etc.), and after about 10 days from contact of somatic cells with reprogramming factors, culture medium for culturing bFGF-containing primate ES cells The ES-like colonies can be generated after about 30 to about 45 days or more from the contact, by culturing in

Alternatively, DMEM culture solution containing 10% FBS on feeder cells (eg, mitomycin C-treated STO cells, SNL cells, etc.) at 37 ° C. in the presence of 5% CO ₂ (in addition, LIF, penicillin / streptomycin, Puromycin, L-glutamine, non-essential amino acids, β-mercaptoethanol, etc. may be included as appropriate) and cultured in contact with somatic cells and reprogramming factor, and after about 25 to about 30 days or more Can give rise to ES-like colonies. Desirably, instead of feeder cells, somatic cells to be reprogrammed are themselves (Takahashi K, et al. (2009), PLoS One. 4: e8067 or WO 2010/137746), or extracellular matrices (eg Laminin- 5 (WO2009 / 123349), Laminin-10 (US 2008/0213885), fragments thereof (WO 2011/043405) and Matrigel (BD) are exemplified. (The documents in this paragraph constitute part of the present application by citation)

  Besides this, a method of establishing iPS cells using a medium not containing serum is also exemplified (Sun N, et al. (2009), Proc Natl Acad Sci USA 106: 15720-15725). Furthermore, to increase establishment efficiency, iPS cells may be established under hypoxic conditions (oxygen concentration of 0.1% or more and 15% or less) (Yoshida Y, et al. (2009), Cell Stem Cell. 5 : 237-241 or WO 2010/013845). (The documents in this paragraph constitute part of the present application by citation)

  Histone deacetylase (HDAC) inhibitor [for example, valproic acid (VPA), trichostatin A, sodium butyrate, small molecule inhibitor such as MC1293, M344, siRNA for HDAC as a component to enhance establishment efficiency of iPS cells And shRNA (eg, nucleic acid expression inhibitors such as HDAC1 siRNA Smartpool (TM) (Millipore), HuSH29mer shRNA Constructs against HDAC1 (OriGene), etc.), MEK inhibitors (eg PD184352, PD98059, U0126, SL327 and PD0325901 ), Glycogen synthetase kinase-3 inhibitors (eg, Bio and CHIR 99021), DNA methyltransferase inhibitors (eg, 5-azacytidine), histone methyltransferase inhibitors (eg, BIX-01294, etc.) Molecular inhibitors, such as nucleic acid expression inhibitors such as siRNA and shRNA against Suv39hl, Suv39h2, SetDBl and G9a) L-channel calcium agonist (eg Bayk 8644), butyric acid, TGFβ inhibitor or ALK5 inhibitor (eg LY364947, SB431542) , 616453 and A-83-01), p53 inhibitors (eg siRNA and shRNA against p53), ARID3A inhibitors (eg siRNA and shRNA against ARID3A), miR-291-3p, miR-294, miR-295 and mir MiRNAs such as -302, Wnt Signaling (eg soluble Wnt3a), Neuropeptide Y, prostaglandins (eg Prostaglandin E2 and Prostaglandin J2), hTER , SV40LT, UTF1, IRX6, GLISl, PITX2, DMRTBl and the like are known. At the time of establishment of iPS cells, a culture solution to which components used for the purpose of improving the establishment efficiency may be used.

During the above culture, culture medium exchange with fresh culture medium is performed once daily from the second day after the culture start. The number of somatic cells used for nuclear reprogramming is not limited, but ranges from about 5 × 10 ³ to about 5 × 10 ⁶ cells per 100 cm ^{2 of} culture dish.

  iPS cells can be selected by the shape of the formed colony. On the other hand, a drug resistant gene expressed in conjunction with a gene (for example, Oct 3/4, Nanog) which is expressed when somatic cells are reprogrammed is introduced as a marker gene, and a culture solution containing a corresponding drug (selected culture solution The established iPS cells can be selected by culturing in In addition, by introducing a fluorescent protein gene as a marker gene and observing with a fluorescent microscope, iPS cells can be selected by adding a luminescent substrate in the case of a luminescent enzyme gene. The induced iPS cells (T-iPS cells) maintain the T cell receptor gene of T cells from which they were derived.

In a second aspect of the present application, a gene of NY-ESO1 antigen specific TCR is introduced into pluripotent stem cells.
For various cancers, Rag1 or Rag2 gene is destroyed by genome editing technology etc. at the stage of NY-ESO1 antigen pluripotent stem cells, and then TCR gene is reported to induce differentiation into T cells. The TCR gene may also be used to isolate desired antigen-specific T cells from cancer patients or derived from healthy individuals, and isolate TCR genes from the T cells. In the present application, a desired antigen-specific TCR gene is introduced into pluripotent stem cells derived from donor cells, such as iPS cells. The introduction of the TCR gene into iPS cells may be carried out by a conventional method, for example, according to the method described in Morgan RA et al, Science, 314: 126. 2006. Specifically, a TCR gene may be placed on an appropriate vector and introduced into iPS cells. For example, it can be introduced into somatic cells by techniques such as viruses, plasmids, vectors such as artificial chromosomes, lipofection, liposomes, microinjection and the like. Examples of viral vectors include retrovirus vectors, lentivirus vectors, adenovirus vectors, adeno-associated virus vectors, Sendai virus vectors and the like. Moreover, examples of artificial chromosome vectors include human artificial chromosome (HAC), yeast artificial chromosome (YAC), bacterial artificial chromosome (BAC, PAC) and the like. As a plasmid, a mammalian cell plasmid can be used. The vector may contain regulatory sequences such as promoter, enhancer, ribosome binding sequence, terminator, polyadenylation site, etc., so that the TCR can be expressed, and further, if necessary, a drug resistance gene (eg, kanamycin resistance). Gene, ampicillin resistance gene, puromycin resistance gene etc.), thymidine kinase gene, selective marker sequences such as diphtheria toxin gene etc., reporter gene sequences such as green fluorescent protein (GFP), β-glucuronidase (GUS), FLAG etc. it can.

  Pluripotent stem cells having a desired TCR gene are induced to differentiate into T precursor cells or mature T cells. Examples of methods for inducing differentiation of pluripotent stem cells into T cells include the methods described in Timmermans et al., Journal of Immunology, 2009, 182: 6879-6888. In addition, in order to prevent the rearrangement of TCR genes before inducing T cells from pluripotent stem cells, Rag1 can be used at the stage of pluripotent stem cells using the technology described in WO 2016/010148. Alternatively, the Rag2 gene may be disrupted by genome editing technology or the like and then induction of differentiation to T cells may be initiated.

  In the present specification and claims, “progenitor cell” refers to the stage of the cell immediately preceding positive selection / negative selection from the stage corresponding to hematopoietic stem cells which are the most undifferentiated cells among hematopoietic cells. Up to the equivalent of Differentiation of T cells is described in Blood 111: 1318 (2008) and Nature Immunology 11: 585 (2010).

  T cells are roughly divided into αβ T cells and γδ T cells, and αβ T cells include killer T cells and helper T cells. The present application is directed to all T cells. T cell includes both T precursor cells and T mature cells, preferably T cells expressing CD3 and expressing at least one molecule selected from the group consisting of CD4 and CD8 It shall be. Preferred are T cells having CTL activity, and CD4 negative CD8 positive cells. T cells (regenerated T cells) containing the same NY-ESO1 antigen-specific TCR by differentiating pluripotent stem cells having the NY-ESO1 antigen-specific TCR into T cells and sorting them appropriately using FACS etc. It is possible to obtain a cell population composed of The purity of the cell population can be further increased by disrupting the Rag1 or Rag2 gene at the stage of pluripotent stem cells by genome editing technology or the like and then inducing differentiation into T cells. For example, by inducing differentiation of iPS cells having NY-ESO1 antigen-specific TCR into T cells, 90% or more, preferably 95% or more of T cells contained in the culture have the same NY-EOS1 specific TCR. T cell cultures can be obtained.

  The expression of NY-ESO1 antigen has been confirmed at a high rate in various types of solid cancer and hematologic malignancies. The present application can be applied to immunocell therapy for cancers that express such NY-ESO1 antigen.

The present application provides T cell preparations that target the NY-ESO1 antigen. As a specific example, NY-ESO1 antigen-specific T cells can be induced from healthy individuals, T-iPS cells can be produced from such cells, and T cells produced from the T-iPS cells can be used. After confirming the function of T cells regenerated from the T-iPS cells using the cells of healthy individuals from which the T-iPS cells are derived, the T-i PS cells are banked and stored.
Examine HLAs of patients suffering from cancer expressing NY-ESO1 antigen to be treated, select compatible HLA-derived cells from banked T-iPS cells, T cells generated from the T-iPS cells Cell immunotherapy is possible. If T-iPS cells are induced to T cells in advance and cryopreserved, they can be administered more rapidly to more patients. Also, since the administered cells are eventually rejected, the risk of canceration of the administered cells may not be considered.

In the immunocell therapy of the present application, the induced T cells are suspended in an appropriate vehicle such as physiological saline or PBS, and administered to a patient to whom HLA matches more than a certain level. When the donor and patient HLA types completely match, when the donor is an HLA haplotype homo, at least one of the HLA haplotypes matches. Administration to patients may be performed intravenously. The dosage is not limited, but 10 ⁶ -10 ⁷ cells / kg in the case of mature T cells, once or more than once, administered intravenously through the patient. In the case of T precursor cells, the dose may be about 1 to 1/1000 of the dose.

The number of cells to be administered is not particularly limited, and may be appropriately determined depending on the age, sex, height, weight, target disease, symptoms and the like of the patient. The optimal number of cells to be administered may be appropriately determined by clinical trials.
T cells can attack various antigens, and the method of the present application can be applied to immune cell therapy for various diseases such as cancer, infections, autoimmune diseases, and allergies.

  The NY-ESO1 gene is highly expressed in melanoma, esophageal cancer, multiple myeloma, adult T-cell leukemia (ATL), etc., and the NY-ESO1 antigen-specific T-i PS cells or TCR-i PS cells to CTL cells When cells induced to differentiate are used, application to immune cell therapy of these various cancers expressing the NY-ESO1 gene is possible.

Generation of iPS cells from NY-ESO1 antigen-specific cytotoxic T lymphocytes (CTL) (NY-ESO1-T-iPS) and regeneration of NY-ESO1 antigen-specific CTL from NY-ESO1-T-iPS

1) Induction and Amplification of NY-ESO1 Antigen-Specific CTL The NY-ESO1 antigen-specific CTL was induced and amplified using a lymphoblastoid cell line LCL (Lymphoblastoid cell line) derived from a healthy individual. The materials used are shown below.
i) Medium

ii) NY-ESO1 peptide SLLMWITQC (SEQ ID NO: 1)
Jaeger E et al. J. Exp. Med. 187: 265-270 (1998)

iii) LCL (Lymphoblastoid cell line)
LCL collected from an HLA-A0201-positive healthy volunteer at Kyoto University Hospital Hematology Oncology (Kyoto City, Kyoto, Japan) was used.

A. Isolation of T cells from human peripheral blood and stimulation with peptide 1. Mononuclear cells were purified by Ficoll from peripheral blood of healthy volunteers and suspended in T cell medium.
2. The cells were seeded at 2.5 × 10 ⁵ / mL per well in a 96-well U-bottom plate, and the peptide was added to 100 nM.
3. T added IL-2 (final concentration 12.5 U / mL), IL-7 (final concentration 5 ng / mL), IL-15 (final concentration 1 ng / mL) on the third day, and added cytokine every week The culture was performed for 2 weeks while changing the medium with the cell culture medium.

B. Peptide addition to LCL LCL was recovered and irradiated with 35 Gy.
2. The cells were suspended in a medium for T cells and adjusted to 5 × 10 ⁵ / mL.
3. The peptide was added to 100 nM and cultured for 2 hours.
4. LCL was recovered, washed with T cell medium, and adjusted to 2 × 10 ⁵ / mL.

C. Co-culture of LCLs pulsed with peptide and T cells After peptide stimulation, T cells cultured for 2 weeks were collected, washed and suspended in a medium for T cells, and adjusted to 2 × 10 ⁶ / mL. At this time, some cells were divided for flow cytometry analysis.
2. 0.5 mL / well of LCL suspension (2 × 10 ⁵ / mL) and 0.5 mL / well of T cell suspension (2 × 10 ⁵ / mL) were cultured in a 24-well plate with added peptide. (LCL: T = 1 × 10 ⁵ / well: 1 × 10 ⁶ / well = 1: 10).
3. On the third day, IL-2 (final concentration 12.5 U / mL), IL-7 (final concentration 5 ng / mL), and IL-15 (final concentration 1 ng / mL) were added. Culture was performed for 2 weeks while changing the medium with T cell medium supplemented with cytokines every week. (Peptide stimulation first course by LCL)
4. LCL was further cultured for 2 hours in a medium supplemented with 100 nM peptide, and CTL was added thereto.
5. On the third day, IL-2 (final concentration 12.5 U / mL), IL-7 (final concentration 5 ng / mL), and IL-15 (final concentration 1 ng / mL) were added. Culture was performed for 2 weeks while changing the medium with T cell medium supplemented with cytokines every week. (2nd course of peptide stimulation by LCL)
6. LCL was further cultured for 2 hours in a medium supplemented with 100 nM peptide, and CTL was added thereto.
7. On the third day, IL-2 (final concentration 12.5 U / mL), IL-7 (final concentration 5 ng / mL), and IL-15 (final concentration 1 ng / mL) were added. Culture was performed for 2 weeks while changing the medium with T cell medium supplemented with cytokines every week. (3rd course of peptide stimulation by LCL)
8. Flow cytometric analysis confirmed that the CD8 positive NY-ESO1 tetramer positive fraction was detected in CD8 positive T cells at a rate of about 10%. The results are shown in FIG.

2) Establishment of NY-ESO1-T-iPS cells Introduction of Yamanaka 4 factor and SV40 by Sendai virus 1. NY-ESO1 tetramer positive CD8 positive cells were sorted by FACS Aria. The sorted NY-ESO1 specific CTL was suspended in a medium for T cells, Sendai virus in which Yamanaka 4 factor and SV40 were incorporated was added to the medium, and the cells were cultured as they were for 2 days.
2 Medium for T cells washed with medium for T cells and further supplemented with IL-2 (final concentration 12.5 U / mL), IL-7 (final concentration 5 ng / mL), IL-15 (final concentration 1 ng / mL) And cultured for 2 days.
3. After all cells were recovered, T cells were suspended in a cytokine-free medium for T cells and seeded onto feeder cells.
4. On the second day, half of the medium for iPS cells was exchanged, and on the next day, half exchange for medium for iPS cells was continued, and culture was continued.

B. iPS cell colonies were visually confirmed 1.3 weeks after pickup of iPS cell colonies.
2. The colonies were physically picked up with a 200 ul tip.
3. Each clone was individually established as iPS cells. The photograph of the obtained clone is shown in FIG.

3) Induction of differentiation of NY-ESO1-T-iPS cells into T cells The medium used was as follows.
* The penicillin / streptomycin solution consisted of penicillin 10000 U / mL and streptomycin 10000 μg / mL, the final concentrations of 100 U / mL and 100 μg / mL respectively.

* The penicillin / streptomycin solution consists of penicillin 10000 U / mL and streptomycin 10000 μg / mL, the final concentrations of 100 U / mL and 100 μg / mL respectively.

Preparation of OP9 Cells 6 ml of a 0.1% gelatin / PBS solution was placed in a 10 cm culture dish and allowed to stand at 37 ° C. for 30 minutes or more. Confluent OP9 cells were detached with a trypsin / EDTA solution, and 1⁄4 equivalent amount was seeded on a gelatin-coated 10 cm culture dish. The medium was added so as to be 10 ml of medium A.
10 ml of medium A was newly added to the OP9 cell culture dish seeded 4 days later to make the total volume 20 ml.

Blood cell precursor cell induction from iPS cells The medium of OP9 cells used for co-culture was aspirated and replaced with fresh medium A. Also, the medium of the iPS cell culture dish was similarly aspirated, and 10 ml of fresh medium A was added. The iPS cell clumps were cut with an EZ-passage roller. The cut iPS cell mass was suspended by pipetting with 200 ul pipetman, and approximately 600 iPS cell masses were visually seeded on OP9 cells.

  Three or more dishes were used per iPS cell clone, and at the time of passaging, the cells were combined once and then redistributed to the same number to reduce the variation among dishes.

Day 1 (medium change)
It was confirmed whether the iPS cell mass had begun to adhere and differentiate, and the medium was replaced with fresh 20 ml of medium A.

Day 5 (half volume change of medium)
Half of the medium was replaced with 10 ml of fresh medium A.

Day 9 (medium change)
Half of the medium was replaced with 10 ml of fresh medium A.

Day 13 (Transfer induced mesodermal cells from OP9 cells onto OP9 / DLL1 cells)
The media was aspirated and the media on the cell surface was flushed with HBSS (+ Mg + Ca). Thereafter, 10 ml of 250 U collagenase IV / HBSS (+ Mg + Ca) solution was added, and the cells were cultured at 37 ° C. for 45 minutes.
The collagenase solution was aspirated and flushed with 10 ml PBS (-). Thereafter, 5 ml of a 0.05% trypsin / EDTA solution was added and cultured at 37 ° C. for 20 minutes. After culture, the cells were peeled off in the form of a membrane, and the cells were physically finely divided by pipetting to separate the adherent cells from each other.

  To this, 20 ml of fresh medium A was added, and culture was further continued at 37 ° C. for 45 minutes. After culture, the supernatant containing floating cells was collected through a 100 μm mesh. The pellet was suspended in 10 ml of medium B by centrifugation at 1200 rpm for 7 minutes at 4 ° C. Among them, 1/10 of them were seeded on freshly prepared OP9 / DLL1 cells, especially for FACS analysis. When cells obtained from a plurality of dishes were pooled, they were redistributed so as to have the same number as the original number, and the cells were replated.

In order to confirm whether the obtained cells contained hematopoietic precursor cells, FACS analysis was performed using an anti-CD34 antibody and an anti-CD43 antibody. Since a sufficient number of cells could be confirmed in the CD34 ^low CD43 ⁺ cell fraction, it was confirmed that hematopoietic progenitor cells were induced.

C. T cell differentiation induction from blood cell precursor cells The cells were then seeded onto OP9 / DLL1 cells. In this step, sorting of the cells of the CD34 ^low CD43 ⁺ cell fraction was not performed. When this fraction is sorted, the efficiency of induction of differentiation into T cells may be lower due to the decrease in the number of cells obtained and the damage to the cells by the sorting than in the case where the sorting is not performed.

  Although FACS analysis is performed to confirm the differentiation stage during the culture period, many dead cells are found in the culture during all periods. Therefore, analysis was performed after removing dead cells using FACS (Propidium Iodide), 7-AAD, etc. at the time of FACS analysis.

Day 16 (passage of cells)
Cells loosely adhering to OP9 cells were gently pipetted several times and collected through a 100 μm mesh into a 50 ml conical tube. The pellet was suspended in 10 ml of medium B by centrifugation at 1200 rpm for 7 minutes at 4 ° C. These cells were seeded on freshly prepared OP9 / DLL1 cells.

Day 23 (passage of cells): Blood cell colonies began to appear.
Cells loosely adhering to OP9 cells were gently pipetted several times and collected through a 100 μm mesh into a 50 ml conical tube. The pellet was suspended in 10 ml of medium B by centrifugation at 1200 rpm for 7 minutes at 4 ° C.

Day 36: Confirmation of NY-ESO1 tetramer positive T cells.
In order to confirm whether or not NY-ESO1 specific T cells were induced, FACS analysis was performed using an anti-CD3 antibody, NY-ESO1 tetramer. The T cell marker CD3 ⁺ cells became visible, and it was confirmed that most of the cells were differentiated to CD3 ⁺ NY-ESO1 tetramer ⁺ . The FACS analysis results are shown in FIG.
Further, when the same cells were further subjected to FACS analysis using anti-CD8 antibody and anti-CD4 antibody, these cells were confirmed to be CD4-CD8 + cells (FIG. 4 left). In addition, when analyzed by an anti-CD8a antibody and anti CD8beta antibody, it was confirmed to have a surface traits of CD8α ⁺ CD8β ⁺ (Fig. 4 right).

Furthermore, the sequences of TCR α chain and β chain of NY-ESO-1 were determined by a conventional method.

The function of the regenerated CTL obtained in Example 1 was evaluated in vitro.
A. Evaluation of peptide-specific cytotoxic activity Whether or not the obtained regenerated CTL has NY-ESO1 peptide-specific CTL activity in vitro is targeted to cells in which NY-ESO-1 peptide antigen is reacted with autologous LCL As, the cytotoxic activity was evaluated.
Autologous LCL was reacted with NY-ESO-1 peptide (1 μM, 100 nM, 10 nM, 1 nM and no peptide (0 nM)) for 2 hours. LCL was recovered, and the regenerated CTL and LCL were mixed at a ratio of 1: 3, 1: 1, 3: 1, 9: 1, and then cultured for 5 hours in an environment of 37 ° C. and 5% CO 2. Cytotoxic activity was then assessed on the percentage of Annexin V positive cells. The results are shown in FIG. As a result, it was shown that the cytotoxic activity was increased depending on the peptide concentration and the number of cells, and it was confirmed that it had NY-ESO-1 antigen specific cytotoxic activity.

B. Evaluation of Cytotoxic Activity on Cell Lines Positive for B A0201 and Expressing NY-ESO1 Adult T-Cell Leukemia Strain TL-Om1 Having A0201 and Expressing NY-ESO-1 and Multiple Myeloma Cell Line U266 The cytotoxic activity of the resulting regenerating CTL was evaluated. As negative controls, NY-ESO-1 positive adult T cell leukemia strains (ATN-1, TL-Su) not expressing A0201 and multiple myeloma strains (RPMI8226, KMS28BM) were used. The results are shown in Figures 6A-C as well as Figures 7A-C.
Regenerated CTL were found to damage A0201-positive and NY-ESO1-expressing cell lines TL-Om1 (FIG. 6A solid line) and U266 (FIG. 7A solid line) in a cell number dependent manner. Moreover, since the cytotoxic activity against TL-Om1 and U266 was suppressed by administering an anti-HLA antibody (FIG. 6A dashed line, FIG. 7A dashed line), it was shown to be an antigen-specific cytotoxic activity. That is, the obtained regenerated CTL recognizes NY-ESO1 in HLA-A0201 restriction and exerts a killer activity.

The function of the regenerated CTL obtained in Example 1 was evaluated in vivo.
It was examined whether the cytotoxic activity of Example 2 was similarly observed in vivo.
Immunodeficient NOG mice were intraperitoneally administered 1 × 10 ⁷ TL-Om-1 cells. Next, PBS was administered to one mouse and 1 × 10 ⁷ regenerated CTLs were administered to the other mouse on the first, second, fourth, ^seventh , ninth, and eleventh disease days, and their survival was analyzed. The results are shown in FIG. In the group to which the regenerated CTL obtained in Example 2 was administered, the survival was significantly prolonged.

Mice were given 1 × 10 ⁷ U266 cells intraperitoneally. Next, PBS was administered to one mouse and 1 × 10 ⁷ regenerated CTL to the other mouse on the first, third, fifth, eighth, fifteenth, and twenty-two disease days, and their survival was analyzed. The results are shown in FIG. In the group to which regenerated CTL was administered, survival was significantly prolonged (FIG. 9). From the above, it was confirmed that the regenerated CTLs also have cytotoxic activity against cancer cells expressing NY-EOS1 even in vivo.

Claims (20)

(1) providing human pluripotent stem cells having NY-ESO1 antigen-specific T cell receptor, and (2) inducing T precursor cells or mature T cells from pluripotent stem cells of step (1). A method of inducing T cells for immune cell therapy, comprising The method according to claim 1, wherein human pluripotent stem cells having NY-ESO1 antigen specific T cell receptor are obtained by deriving pluripotent stem cells from NY-ESO1 antigen specific human T cells. NY-ESO1 antigen-specific human T cells are not immunocytotherapy subjects but T cells obtained from humans who have or have a history of having a cancer that expresses NY-ESO1 antigen, The method of claim 2. The NY-ESO1 antigen-specific human T cell is not an immune cell therapy subject but a T cell obtained from a human who has not suffered from a cancer that expresses the NY-ESO1 antigen. Method. A human pluripotent stem cell having a NY-ESO1 antigen-specific T cell receptor is a pluripotent stem cell obtained by introducing a NY-ESO1 antigen-specific T cell receptor gene into human pluripotent stem cells The method according to claim 1. The method according to any one of claims 1 to 5, wherein the human pluripotent stem cell having NY-ESO1 antigen-specific T cell receptor is one in which Rag2 or Rag2 gene is deleted. The human pluripotent stem cell having an antigen-specific T cell receptor is a human-derived pluripotent stem cell having an HLA type that completely or partially matches the HLA type of a subject of immune cell therapy. The method in any one of 1-6. Human pluripotent stem cells having an antigen-specific T cell receptor can be obtained from a human having a haplotype homozygous HLA having one or more haplotypes of the HLA of the immuno cell therapy subject. The method according to claim 7, which is a T cell. The method according to any one of claims 1 to 8, wherein the human pluripotent stem cells are human iPS cells. The method according to any one of claims 1 to 9, wherein the immune cell therapy is for treating cancer. 11. The method according to claim 10, wherein the cancer is a cancer that expresses the NY-ESO1 gene. The method according to claim 11, wherein the cancer is selected from the group consisting of melanoma, esophageal cancer, multiple myeloma, adult T cell leukemia (ATL). The method according to any one of claims 1 to 12, wherein the NY-ESO1 antigen-specific T cell receptor recognizes the peptide SLLMWITQC in an HLA-A0201 restricted manner. The method according to any one of claims 1 to 13, wherein the NY-ESO1 antigen-specific T cell receptor has CAVNTDSNYQLIW in the TCR α chain and CASSVAAGE QFF in the TCR β chain as the amino acid sequence of the antigen recognition site. NY-ESO1 antigen-specific T cell receptor for immune cell therapy iPS cells. The immunoPSA iPS cell according to claim 15, wherein the NY-ESO1 antigen-specific T cell receptor recognizes the peptide SLLMWITQC under HLA-A0201 restriction. 16. The iPS cell for immune cell therapy according to claim 15, wherein the NY-ESO1 antigen-specific T cell receptor has, as an amino acid sequence of an antigen recognition site, CAVNTDSNYQLIW in the TCR α chain and CASSVAAGE QFF in the TCR β chain. An immune cell therapy T cell culture comprising 90% or more of T cells having the same NY-ESO1 antigen-specific T cell receptor. The T cell culture for immune cell therapy according to claim 18, wherein the NY-ESO1 antigen-specific T cell receptor recognizes the peptide SLLMWITQC in an HLA-A0201 restricted manner. The immunocytotherapy T-cell culture according to claim 18, wherein the NY-ESO1 antigen-specific T cell receptor has CAVNTDSNYQLIW in the TCR α chain and CASSVAAGE QFF in the TCR β chain as the amino acid sequence of the antigen recognition site.