This document relates to bioengineering and involves bioengineered thymus organoids and related humanized animal models. The thymus organoids and animal models have various commercial and clinical uses, including generating humanized antibodies, making antigen-specific human T cells, inducing transplantation tolerance, rejuvenating thymus functions, and modeling human diseases.

The present description provides improved methods for generating thymic epithelial progenitor (TEP) cells from pluripotent stem (PS) cells in vitro. Also provided are isolated invitro cell populations, compositions, and systems comprising TEP cells produced in vitro. Compositions and systems of cell populations of thymic epithelial cells and subpopulations thereof, as well as cells formed during different stages of differentiation of PS cells into thymic epithelial cells and subpopulations thereof are provided.

In some embodiments, compositions and methods re¬lating to isolated artificial antigen presenting cells (aAPCs) are dis¬closed, including aAPCs comprising a myeloid cell transduced with one or more viral vectors, such as a MOLM-14 or a EM-3 myeloid cell, wherein the myeloid cell endogenously expresses HLA-A/B/C, ICOS-L, and CD58, and wherein the one or more viral vectors com¬prise a nucleic acid encoding CD86 and a nucleic acid encoding 4-1BBL and/or OX40L and transduce the myeloid cell to express CD86 and 4-1BBL and/or OX40L proteins. In some embodiments, methods of expanding tumor infiltrating lymphocytes (TILs) with aAPCs and methods of treating cancers using TILs after expansion with aAPCs are also disclosed.

Methods for generating thymic epithelial progenitor (TEP) cells from pluripotent stem (PS) cells in vitro are provided. Compositions and systems of cell populations of TEP cells as well as cells formed during different stages of differentiation of PS cells into TEP cells are also disclosed. The methods, isolated in vitro cell populations, compositions, and systems disclosed provide functional TEP cells that mature into thymic epithelial cells in vivo.

In some embodiments, compositions and methods relating to isolated artificial antigen presenting cells (aAPCs) are disclosed, including aAPCs comprising a myeloid cell transduced with one or more viral vectors, such as a MOLM-14 or a EM-3 myeloid cell, wherein the myeloid cell endogenously expresses HLA-AB/C, ICOS-L, and CD58, and wherein the one or more viral vectors comprise a nucleic acid encoding CD86 and a nucleic acid encoding 4-1BBL and/or OX40L and transduce the myeloid cell to express CD86 and 4-1BBL and/or OX40L proteins. In some embodiments, methods of expanding tumor infiltrating lymphocytes (TILs) with aAPCs and methods of treating cancers using TILs after expansion with aAPCs are also disclosed.

The current disclosure provides for methods of promoting differentiation of pluripotent stem cells into thymic epithelial cells or thymic epithelial cell progenitors as well as the cells obtained from the methods, and solutions, compositions, and pharmaceutical compositions comprising such cells. The current disclosure also provides for methods of using the thymic epithelial cells or thymic epithelial cell progenitors for treatment and prevention of disease, generating organs, as well as other uses, and kits.

Embodiments disclosed herein concern various cells, co-cultures, methods, systems, therapies, and treatments involving in vitro and in vivo generation and use of functional mammalian thymic tissue, thymus organs, and thymic organoids. In certain embodiments, cells, tissues, and organoids disclosed herein can be used to treat a subject having a thymic condition. In other embodiments, thymic cells and organoids produced by compositions and methods disclosed herein can be used to treat various conditions, diseases, and disorders, including auto-immune disorders, transplant rejections, cancer, or aging.

In some embodiments, compositions and methods relating to isolated artificial antigen presenting cells (aAPCs) are disclosed, including aAPCs comprising a myeloid cell transduced with one or more viral vectors, such as a MOLM-14 or a EM-3 myeloid cell, wherein the myeloid cell endogenously expresses HLA-A/B/C, ICOS-L, and CD58, and wherein the one or more viral vectors comprise a nucleic acid encoding CD86 and a nucleic acid encoding 4-1BBL and/or OX40L and transduce the myeloid cell to express CD86 and 4-1BBL and/or OX40L proteins. In some embodiments, methods of expanding tumor infiltrating lymphocytes (TILs) with aAPCs and methods of treating cancers using TILs after expansion with aAPCs are also disclosed.

Methods and compositions for promoting donor-specific tolerance and immunocompetence to a recipient of a solid organ transplant, by implanting an allogeneic solid organ in a recipient in need of a solid organ transplant and further comprising surgical implantation of a tissue-engineered allogeneic cultured postnatal thymus tissue product in the recipient of a solid organ from a donor.

The disclosed technology includes methods, systems, and devices for generating patient-specific functional thymic epithelial progenitor (TEP) cells. In some implementations, a method may include generating iPSCs from HSC; causing differentiation of the iPSC into thymic epithelial progenitor (TEP) cells, generating thymic epithelial cells by transplantation of the TEP cells into a host, wherein the TEP cells may differentiate into mature functional thymic epithelial cells (TECs). In some implementations, a system may include a cell population of patient specific cells, a population of iPSCs, a culture system for differentiating the iPSCs into a population of patient-specific TEP cells for transfer to a host or the patient to allow the TEP cells to differentiate into mature, functional TEC.