# Generation of thymus-reconstituting T cell progenitors from human pluripotent stem cells

## TL;DR

Scientists created T cell progenitors from human stem cells that can regenerate the thymus and mature into functional T cells.

## Contribution

A protocol for generating thymus-reconstituting T cell progenitors from human pluripotent stem cells is developed.

## Key findings

- hPSC-derived pro-T cells resemble early thymic progenitors in transcription profiles.
- The generated pro-T cells reconstitute the thymus and mature into T cells in vivo.
- The cells have a specific surface marker profile (CD4‒/lowCD8‒CD7+CD34+CD45RA+).

## Abstract

Generating a large number of progenitors that can repopulate the immune system of a recipient is one of the key steps toward efficient cancer immunotherapy. Here, we describe the engineering of T cell progenitors capable of direct and long-term reconstitution of the thymus. In the thymus, human pluripotent stem cell (hPSC)-derived progenitor T cells (pro-T cells) developed into single-positive human T cells that entered circulation and settled in the spleen. Single-cell transcriptome analysis of differentiating hPSCs attested to the emergence of cells that displayed the transcription signature of the early T cell progenitors. Comparative transcription profiling revealed the similarity of the hPSC-pro-T cells with the early T cell precursors of the human thymus. The in vitro generation of T cell progenitors provides a powerful model for studying the molecular mechanisms of human T cell development and improves the perspectives for T cell regenerative medicine, including chimeric antigen receptor T (CAR-T) cell therapies.

•We establish a protocol for generation of repopulating hPSC-derived T cell progenitors•Transcriptome profiles of the hPSC-pro-T cells are similar to early thymic progenitors•The hPSC-derived pro-T cells have the CD4‒/lowCD8‒CD7+CD34+CD45RA+ phenotype•The hPSC-pro-T cells reconstitute the thymus and develop in vivo into mature T cells

We establish a protocol for generation of repopulating hPSC-derived T cell progenitors

Transcriptome profiles of the hPSC-pro-T cells are similar to early thymic progenitors

The hPSC-derived pro-T cells have the CD4‒/lowCD8‒CD7+CD34+CD45RA+ phenotype

The hPSC-pro-T cells reconstitute the thymus and develop in vivo into mature T cells

Fighting cancer and immunodeficiency disorders by adoptive transfer of T cell progenitors has certain advantages over hematopoietic stem cell transplantations (HSCTs). De novo generation of donor-HSC-derived T cells is slowed down at bone marrow (BM) seeding, the generation of T cell progenitors in BM, and delivery to the thymus. Lagging thymus reconstitution upon HSCT would lead to an extended period of immunodeficiency with increased health risks. Finding the source of autologous thymus-reconstituting pro-T cells would be instrumental in alleviating the T cell recovery delay upon HSCT. To address this issue, we developed a protocol for generating transplantable pro-T cells from human pluripotent stem cells.

Philonenko et al. report generation of T cell progenitors from human pluripotent stem cells (hPSCs). These hPSC-pro-T cells demonstrate the single-cell transcription profile of early thymic progenitors from the human fetus. The T cell progenitors reconstitute the thymus over the long term and develop into mature T cells in vivo.