Lineage tracing of T cell differentiation from T-iPSC by 2D feeder-free culture and 3D organoid culture

Yoshitaka Ishiguro; Shoichi Iriguchi; Shinya Asano; Tokuyuki Shinohara; Sara Shiina; Suguru Arima; Yoshiaki Kassai; Yoshiharu Sakai; Kazutaka Obama; Shin Kaneko

doi:10.3389/fimmu.2023.1303713

Lineage tracing of T cell differentiation from T-iPSC by 2D feeder-free culture and 3D organoid culture

Front Immunol. 2023 Dec 15:14:1303713. doi: 10.3389/fimmu.2023.1303713. eCollection 2023.

Authors

Yoshitaka Ishiguro^{1

2

3}, Shoichi Iriguchi^{1

2}, Shinya Asano⁴, Tokuyuki Shinohara^{2

5}, Sara Shiina^{1

2}, Suguru Arima^{2

5}, Yoshiaki Kassai^{2

5}, Yoshiharu Sakai⁶, Kazutaka Obama³, Shin Kaneko^{1

2}

Affiliations

¹ Shin Kaneko Laboratory, Department of Cell Growth and Differentiation, Kyoto University, Shogoin-Kawahara-cho, Sakyo-ku, Kyoto, Japan.
² Takeda-CiRA Joint Program (T-CiRA), Fujisawa, Japan.
³ Department of Surgery, Graduate School of Medicine, Kyoto University, Shogoin-Kawahara-cho, Sakyo-ku, Kyoto, Japan.
⁴ Axcelead Drug Discovery Partners, Inc., Fujisawa, Japan.
⁵ T-CiRA Discovery and Innovation, Takeda Pharmaceutical Company, Fujisawa, Japan.
⁶ Department of Surgery, Osaka Red Cross Hospital, Fudegasaki-cho, Tennoji-ku, Osaka, Japan.

Abstract

Introduction: T cells induced from induced pluripotent stem cells(iPSCs) derived from antigen-specific T cells (T-iPS-T cells) are an attractive tool for T cell immunotherapy. The induction of cytotoxic T-iPS-T cells is well established in feeder-free condition for the aim of off-the-shelf production, however, the induction of helper T-iPS-T cells remains challenging.

Methods: We analyzed T-iPS-T cells matured in 3D organoid culture at different steps in the culture process at the single-cell level. T-iPS-T cell datasets were merged with an available human thymocyte dataset based in single-cell RNA sequencing (scRNA-seq). Particularly, we searched for genes crucial for generation CD4+ T-iPS-T cells by comparing T-iPS-T cells established in 2D feeder-free or 3D organoid culture.

Results: The scRNA-seq data indicated that T-iPS-T cells are similar to T cells transitioning to human thymocytes, with SELENOW, GIMAP4, 7, SATB1, SALMF1, IL7R, SYTL2, S100A11, STAT1, IFITM1, LZTFL1 and SOX4 identified as candidate genes for the 2D feeder-free induction of CD4+ T-iPS-T cells.

Discussion: This study provides single cell transcriptome datasets of iPS-T cells and leads to further analysis for CD4+ T cell generation from T-iPSCs.

Keywords: 3D organoid; CD4; T cell differentiation; iPSC; scRNA seq.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cell Culture Techniques
Cell Differentiation
GTP-Binding Proteins
Genes, Homeobox
Humans
Induced Pluripotent Stem Cells*
Matrix Attachment Region Binding Proteins*
Organoids
SOXC Transcription Factors

Substances

SATB1 protein, human
Matrix Attachment Region Binding Proteins
SOX4 protein, human
SOXC Transcription Factors
GIMAP4 protein, human
GTP-Binding Proteins

Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was supported by the Interuniversity Bio-Backup Project for Basic Biology (IBBP). The entire study was conducted in accordance with the Declaration of Helsinki and permitted by the institutional ethical board of Kyoto University. This work was supported in part by the Ministry of Education, Culture, Sports, Science, and Technology of Japan (23591413, 15H04655, 15J05263, 26293357), Japan Agency for Medical Research and Development (Project for Development of Innovative Research on Cancer Therapeutics, Practical Research for Innovative Cancer Control, and Core Center for iPS Cell Research), and the Takeda-CiRA collaboration program.