Advancing iPSC-derived Thymic Epithelial Cells as Cell Therapy for T Cell Immune Reconstitution in Vulnerable Populations(original application ID AI190181-01)

| NARRATIVE The thymus instructs T cell immunity and central tolerance, yet its therapeutic potential remains clinically untapped as the signals that drive thymic epithelial cell (TEC) differentiation remain incompletely understood. The thymic epithelium comprises a highly specialized set of cells that attract lymphoid progenitors, promote their proliferation and maturation into thymocytes, and facilitate the selection of a diverse, self-tolerant T cell receptor (TCR) repertoire. The role of the thymus in building immune identity begins before birth. The organ peaks in size in infancy and then structurally and functionally involutes over time. This process causes the decline in immune competence with age (immune senescence). The impact of this phenomenon was exposed during the COVID-19 pandemic when waning immunity left the elderly more vulnerable to adverse outcomes. Thymus insult also occurs in many patients through medications, radiation, infections and graft-versus-host disease. The most severe form of thymic compromise is congenital athymia, the inborn absence of the thymus due to genetic mutations. Genetic or acquired thymic injury leads to immunodeficiency, autoimmunity, inflammation and increased cancer risk. Regenerating thymic function, e.g., through human induced pluripotent stem cell (iPSC)-derived regenerative thymic tissues holds greatest therapeutic promise for these patients. We have used single-cell transcriptomics of human fetal anterior foregut-derived organs to uncover the signals that drive TEC differentiation. We have translated these insights into a novel differentiation platform for the derivation of TECs from iPSCs in vitro. When iPSC-derived TEC organoids are transplanted into athymic NSG nude (NSG-Foxn1-/-) mice engrafted with human hematopoietic stem cells, they function like the human thymus, giving rise to human ab-T cells with a diverse TCR repertoire, gd-T cells and regulatory T cells. In this application, we now seek to advance the translation of iPSC-derived TECs (iTECs) by testing their safety and efficacy as cell therapy for vulnerable patient populations in need of improved T cell immunity. In Aim 1, we will determine the capacity of iTECs to promote T cell reconstitution, functional antigen-specific T cell responses and the development of a broad TCR repertoire in vivo. In Aim 2, we will assess if T cells educated on iTEC are tolerant to “self” but respond to “non-self”. In addition, we will directly analyze the HLA-associated peptide repertoire presented on iTECs using immunopeptidomics. In Exploratory Aim 3, we will test if HLA-editing of iPSCs for iTECs derivation affects antigen-specific immune responses, TCR repertoire, and immunopeptidome in vivo. Advancing the translation of iPSC-derived TECs into a cell therapy is an entirely new strategy to leverage the therapeutic potential of T cells from inside the body and could begin a new chapter of immunotherapeutics. Project Number: 1R01AI190181-01A1 | Fiscal Year: 2026 | NIH Institute/Center: National Institute of Allergy and Infectious Diseases (NIAID) | Principal Investigator: Katja Weinacht | Institution: STANFORD UNIVERSITY, STANFORD, CA | Award Amount: $809,250 | Activity Code: R01 | Study Section: Advancing Therapeutics - B Study Section[ATB] View on NIH RePORTER: https://reporter.nih.gov/project-details/1R01AI19018101A1