Projects

The present project aims at producing “universal” T cells that can be given to any patient. While such universal T cells can be used for various diseases, we plan to apply this project initially to cancer patient (Figure 1). As a material of such T cells, we are going to produce universal ES cells or iPS cells.
It has been shown that the adoptive T cell therapy is effective for some types of cancer. The currently ongoing adoptive T cell therapies have been conducted in an autologous setting; T cells collected from a patient are transferred back to the patient after genetic modification and expansion. However, such methods have faced some problems: these methods are costly, time-consuming, and unstable in quality since they depend on the patient’ T cells. To address these issues, we have developed a method to mass-produce T cells, which will make it possible to prepare T cells that can be used in an allogeneic setting; in other words, to prepare universal T cells that can be given to anyone.
To this end, in 2013, we succeeded in producing iPS cells from T cells and to regenerate T cells from such iPS cells (T-iPSC method) (Figure 2). With this method, it becomes possible to mass-produce T cells having a certain antigen specificity. Subsequently, we developed a method to transduce iPS cells with exogenous T cell receptor (TCR) gene and to regenerate T cells from such iPS cells (TCR-iPSC method) (Figure 3). With this method, it has become possible to use ES cells as well as iPS cells.
Then, what kind of ES/iPS cells should be used as a cell source? When your cells are transferred to another person, such cells will be rejected soon unless HLA type is matched between you and the recipient. At present, in the regenerative medicine field, a major project has been promoted by Japanese government, in which HLA haplotype-homozygous iPS cells are banked. In this strategy, it is expected that cells/tissues regenerated from such iPS cells can be transplanted to patients who retain the same HLA haplotype on one allele, because immune system in such patients may make minimal immune response against the graft. While this strategy would work well, some concern remains; even when you prepare as many as top 10 frequent iPS cell lines, they can cover only 50 % of Japanese people. Moreover, our own research has revealed that the graft will be rejected by NK cells at a certain frequency (2017).
To address such issue, in the present project, we are going to develop “universal” T cells that can be transfused to any patients. As a cell source for such T cells, we will firstly produce universal pluripotent stem cells. To this end, HLA will be genetically deleted in the stem cells as a basic strategy. In such a case, it is expected that NK cell-mediated immune reaction takes place. Thus, one of main aims of this project is to develop a method that can cancel such immune reaction. As a cell source, we will mainly use ES cells, while we will also use iPS cells.
One of other concerns in the HLA-deleted pluripotent stem cells is that the regenerated cells are susceptible to infection by virus or bacteria in patient’s body. In the present project, we will develop a method to solve this issue.
Very recently, just prior to this project, we have succeeded in improving the TCR-iPSC method, making it possible to “efficiently” and “safely” transduce ES/iPS cells with exogenous TCR gene; to firstly knock-in a cassette deck structure into ES/iPS cells and then to insert TCR gene as a cassette tape (Figure 4). In the present project, we will further improve this method.
To evaluate function of the regenerated T cells, we are going to use PDX (patient-derived xenograft) model, in which tumor tissue collected from a patient is transplanted in immunodeficient mouse. One point here is that the universal T cells are expected to work in the presence of white blood cells of the patient. Therefore, we plan to establish PDX model in which peripheral blood mononuclear cells from the same patient are also transfused. This project will be conducted mainly by Kyoto University, in collaboration with Osaka University, Shiga University of Medical Science, and Riken IMS (Figure 5).

Figure 1: The goal of the present project
We firstly produce universal pluripotent stem cells (ES cells or iPS cells). Then, T cells that can attack cancer cells are mass-produced from such ES/iPS cells. Such T cells will be frozen and banked as “off-the-shelf T cells”. When needed, T cells will be thawed and intravenously transfused to a cancer patient. The transfused T cells will attack cancer cells while immune cells of the patient will not attack them.

Figure 2: Regeneration of killer T cells specific for tumor antigen
Mart1 antigen-specific killer T cells derived from a melanoma patient were reprogrammed into iPS cells (Cell Stem Cell, 12: 31, 2013). Killer T cells regenerated from such iPS cells were found to express the same T cell receptor as the original T cells.

Figure 3: TCR-iPSC method
T cells are produced from iPS cells that have been transduced with exogenous TCR gene. Merits of this method are: 1) possible to use high quality iPS cells provided by Center for iPS cell Research and Application (Kyoto University), 2) possible to use TCR gene efficacy and safety of which have been clinically tested.

Figure 4: TCR cassette method
A cassette deck structure is knocked-in into genome of ES/iPS cells. TCR gene is inserted into the cassette deck site as a cassette tape using the RMCE（recombinase mediated cassette exchange）method.

Figure 5: Framework of the project
Hiroshi Kawamoto (Kyoto University) leads the project in collaboration with other members in Kyoto University consisting of Masahiro Inoue, Toshio Kitawaki, Masaki Mandai, and Junzo Hamanishi, and with members of other organizations consisting of Hisashi Arase (Osaka University), Yasutoshi Agata (Shiga University of Medical Science) and Ichiro Taniuchi (Riken IMS).