scholarly journals Keratinocyte growth factor augments immune reconstitution after autologous hematopoietic progenitor cell transplantation in rhesus macaques.

Blood ◽  
2007 ◽  
Vol 110 (1) ◽  
pp. 441-449 ◽  
Author(s):  
Ruth Seggewiss ◽  
Karin Loré ◽  
F. Javier Guenaga ◽  
Stefania Pittaluga ◽  
Joseph Mattapallil ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Growth Factor ◽  
Immune Reconstitution ◽  
Progenitor Cell ◽  
Rhesus Macaques ◽  
Keratinocyte Growth Factor ◽  
Hematopoietic Progenitor Cell ◽  
Thymic Epithelial Cells ◽  
T Cell Reconstitution

Opportunistic infections contribute to morbidity and mortality after peripheral blood progenitor cell (PBPC) transplantation and are related to a deficient T-cell compartment. Accelerated T-cell reconstitution may therefore be clinically beneficent. Keratinocyte growth factor (KGF) has been shown to protect thymic epithelial cells in mice. Here, we evaluated immune reconstitution after autologous CD34+ PBPC transplantation in rhesus macaques conditioned with myeloablative total body irradiation in the absence or presence of single pretotal body irradiation or repeated peritransplant KGF administration. All KGF-treated animals exhibited a well-preserved thymic architecture 12 months after graft. In contrast, thymic atrophy was observed in the majority of animals in the control group. The KGF-treated animals showed higher frequencies of naive T cells in lymph nodes after transplantation compared with the control animals. The animals given repeated doses of KGF showed the highest levels of T-cell receptor excision circles (TRECs) and the lowest frequencies of Ki67+ T cells, which suggest increased thymic-dependent reconstitution in these animals. Of note, the humoral response to a T-cell–dependent neo-antigen was significantly higher in the KGF-treated animals compared with the control animals. Thus, our findings suggest that KGF may be a useful adjuvant therapy to augment T-cell reconstitution after human PBPC transplantation.

Keratinocyte growth factor preserves normal thymopoiesis and thymic microenvironment during experimental graft-versus-host disease

Blood ◽  
2002 ◽  
Vol 100 (2) ◽  
pp. 682-691 ◽  
Author(s):  
Simona Rossi ◽  
Bruce R. Blazar ◽  
Catherine L. Farrell ◽  
Dimitry M. Danilenko ◽  
David L. Lacey ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Growth Factor ◽  
Graft Versus Host Disease ◽  
Keratinocyte Growth Factor ◽  
Receptor Expression ◽  
Successful Outcome ◽  
Thymic Epithelial Cells ◽  
Host Disease ◽  
Graft Versus Host

Abstract Thymus-dependent reconstitution of the peripheral T-cell compartment is critical for the successful outcome of bone marrow transplantation. However, graft-versus-host disease (GVHD) affects thymic stromal function and thus prevents normal T-cell maturation and selection. To determine whether cytoprotection of thymic epithelial cells (TECs) by keratinocyte growth factor (KGF) averts GVHD-related injury to the thymus, a nonirradiated murine parent→F1 transplantation model was investigated. Administration of KGF between days −3 and +3 of GVHD induction preserved normal thymic size, cellularity, and thymocyte phenotype when measured 2 weeks after transplantation and compared with saline-treated parent→F1 mice that received allogeneic transplants. Moreover, the characteristic GVHD-induced impairment in cell cycle progression of pro- and pre-T cells was prevented by KGF. However, the normal phenotypic and functional status of the thymus did not correlate with the higher number of GVHD-inducing mature donor T cells in thymi of KGF-treated mice. Importantly, extensive analysis of the different TEC populations within the thymic cortex and medulla revealed an almost normal stromal architecture and composition in GVHD mice treated with KGF. These observations are likely to reflect an indirect effect of KGF on thymopoiesis as KGF-receptor expression was demonstrated to be restricted to TECs. Thus, pharmacologic doses of KGF appear to exert a potent effect on TEC function, which in turn allows for normal T lymphopoiesis to occur during acute GVHD.

scholarly journals Keratinocyte growth factor enhances DNA plasmid tumor vaccine responses after murine allogeneic bone marrow transplantation

Blood ◽  
2009 ◽  
Vol 113 (7) ◽  
pp. 1574-1580 ◽  
Author(s):  
Robert R. Jenq ◽  
Christopher G. King ◽  
Christine Volk ◽  
David Suh ◽  
Odette M. Smith ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Keratinocyte Growth Factor ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Effector Cells ◽  
Cd8 Cells ◽  
T Cell Reconstitution ◽  
Dna Plasmid

Abstract Keratinocyte growth factor (KGF), which is given exogenously to allogeneic bone marrow transplantation (allo-BMT) recipients, supports thymic epithelial cells and increases thymic output of naive T cells. Here, we demonstrate that this improved T-cell reconstitution leads to enhanced responses to DNA plasmid tumor vaccination. Tumor-bearing mice treated with KGF and DNA vaccination have improved long-term survival and decreased tumor burden after allo-BMT. When assayed before vaccination, KGF-treated allo-BMT recipients have increased numbers of peripheral T cells, including CD8+ T cells with vaccine-recognition potential. In response to vaccination, KGF-treated allo-BMT recipients, compared with control subjects, generate increased numbers of tumor-specific CD8+ cells, as well as increased numbers of CD8+ cells producing interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α). We also found unanticipated benefits to antitumor immunity with the administration of KGF. KGF-treated allo-BMT recipients have an improved ratio of T effector cells to regulatory T cells, a larger fraction of effector cells that display a central memory phenotype, and effector cells that are derived from a broader T-cell–receptor repertoire. In conclusion, our data suggest that KGF can function as a potent vaccine adjuvant after allo-BMT through its effects on posttransplantation T-cell reconstitution.

Keratinocyte growth factor facilitates alloengraftment and ameliorates graft-versus-host disease in mice by a mechanism independent of repair of conditioning-induced tissue injury

Blood ◽  
2000 ◽  
Vol 96 (13) ◽  
pp. 4350-4356 ◽  
Author(s):  
Angela Panoskaltsis-Mortari ◽  
Patricia A. Taylor ◽  
Jeffrey S. Rubin ◽  
Aykut Uren ◽  
Lisbeth A. Welniak ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Growth Factor ◽  
Graft Versus Host Disease ◽  
Keratinocyte Growth Factor ◽  
Allogeneic Bone ◽  
Cell Transfer ◽  
Host Disease ◽  
Graft Versus Host ◽  
T Cell Transfer

Abstract We have previously shown that pretreatment of mice with keratinocyte growth factor (KGF), an epithelial tissue repair factor, can ameliorate graft-versus-host disease (GVHD) after intensive chemoradiotherapeutic conditioning and allogeneic bone marrow transplantation (BMT). To determine whether this effect was dependent on a KGF-mediated mechanism affecting repair of conditioning-induced epithelial cell injury, we studied GVHD in the absence of conditioning using BALB/c severe combined immune-deficient (SCID) recipients given C57BL/6 T cells. KGF (5 mg/kg per day, subcutaneously) given either before or after T-cell transfer enhanced body weights and extended survival. KGF-treated recipients had elevated serum levels of the Th2 cytokine interleukin 13 (IL-13) on day 6 after T-cell transfer concomitant with reduced levels of the inflammatory cytokines tumor necrosis factor-α (TNF-α) and interferon gamma (IFN-γ). A 3-day KGF pretreatment also depressed the secondary in vitro mixed lymphocyte response (MLR) of C57BL/6 splenocytes taken 7 days after in vivo alloimmunization with irradiated BALB/c spleen cells. To determine whether KGF would inhibit host-antidonor–mediated BM rejection, pan-T-cell–depleted BALB/c BM cells were infused into sublethally irradiated C57BL/6 mice and administered KGF either before or before and after BMT. Surprisingly, all KGF schedules tested actually resulted in enhanced alloengraftment. The presence of KGF receptor on donor antihost alloreactive T cells could not be detected by binding studies with radiolabeled KGF, reverse transcriptase–polymerase chain reaction, and Western blotting. Therefore, the mechanism of action of KGF on inhibiting T-cell–mediated immune effects may not be due to a direct effect of KGF on T cells. These studies demonstrate that KGF, by mechanisms independent of repair of conditioning-induced injury, has great potential as an anti-GVHD therapeutic agent with the added benefit of inhibiting the rejection of pan-T-cell–depleted donor BM allografts.

scholarly journals Short-term inhibition of p53 combined with keratinocyte growth factor improves thymic epithelial cell recovery and enhances T-cell reconstitution after murine bone marrow transplantation

Blood ◽  
2010 ◽  
Vol 115 (5) ◽  
pp. 1088-1097 ◽  
Author(s):  
Ryan M. Kelly ◽  
Emily M. Goren ◽  
Patricia A. Taylor ◽  
Scott N. Mueller ◽  
Heather E. Stefanski ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Transplantation ◽  
T Cell ◽  
Growth Factor ◽  
Epithelial Cell ◽  
Marrow Transplantation ◽  
Keratinocyte Growth Factor ◽  
Thymic Epithelial Cell ◽  
Cell Recovery ◽  
T Cell Reconstitution

Abstract Myeloablative conditioning before bone marrow transplantation (BMT) results in thymic epithelial cell (TEC) injury, T-cell immune deficiency, and susceptibility to opportunistic infections. Conditioning regimen–induced TEC damage directly contributes to slow thymopoietic recovery after BMT. Keratinocyte growth factor (KGF) is a TEC mitogen that stimulates proliferation and, when given before conditioning, reduces TEC injury. Some TEC subsets are refractory to KGF and functional T-cell responses are not fully restored in KGF-treated BM transplant recipients. Therefore, we investigated whether the addition of a pharmacologic inhibitor, PFT-β, to transiently inhibit p53 during radiotherapy could spare TECs from radiation-induced damage in congenic and allogeneic BMTs. Combined before BMT KGF + PFT-β administration additively restored numbers of cortical and medullary TECs and improved thymic function after BMT, resulting in higher numbers of donor-derived, naive peripheral CD4+ and CD8+ T cells. Radiation conditioning caused a loss of T-cell zone fibroblastic reticular cells (FRCs) and CCL21 expression in lymphoid stroma. KGF + PFT-β treatment restored both FRC and CCL21 expression, findings that correlated with improved T-cell reconstitution and an enhanced immune response against Listeria monocytogenes infection. Thus, transient p53 inhibition combined with KGF represents a novel and potentially translatable approach to promote rapid and durable thymic and peripheral T-cell recovery after BMT.

Reactivity of Resident Immunocytes in Normal and Prepsoriatic Skin Using an Ex Vivo Skin-Explant Model System

2003 ◽  
Vol 127 (3) ◽  
pp. 289-296 ◽  
Author(s):  
Jonathan L. Curry ◽  
Jian-Zhong Qin ◽  
June Robinson ◽  
Brian J. Nickoloff
Keyword(s):  
T Cells ◽  
T Cell ◽  
Growth Factor ◽  
Ex Vivo ◽  
Keratinocyte Growth Factor ◽  
Experimental System ◽  
Interferon Γ ◽  
T Cell Subsets ◽  
Trigger Factors ◽  
Molecular Events

Abstract Context.—While it is well known that both exogenous and endogenous stimuli can trigger appearance of psoriatic lesions, the initial cellular and molecular events mediated by immunocompetent cells normally resident in prepsoriatic (PN) skin are not well understood. Moreover, it is unclear whether there are any fundamentally important differences in the innate immune response of normal healthy skin (NN skin) versus PN skin. Since acute tissue responses to stimuli involve both resident cells and immunocytes recruited rapidly from circulation, it is difficult to discern the contribution of endogenous cells normally present in skin. Objective.—To solely characterize the reactivity of resident immunocytes using an experimental system. Design.—To probe the activation potential of resident immunocytes in NN (n = 18) and PN skin (n = 10), a short-term ex vivo organ culture system containing interleukin (IL)-2 was established and characterized. To mimic exogenous or environmental trigger factors, bacteria-derived superantigens and lipopolysaccharide were added to the skin-explant assays, whereas endogenous trigger factors were investigated using heat shock proteins. Results.—Using this skin-explant assay, both NN and PN skin gave rise to an expansion of various T-cell subsets, which could differentially produce various cytokines and a growth factor (keratinocyte growth factor), depending on the stimulus and source of skin. Bacterial superantigens were relatively potent inducers of interferon-γ, and natural killer–T cells were observed proliferating from PN skin. Conclusions.—Despite relatively few T cells normally residing in either NN or PN skin, initiation of skin explants from both sets of individuals in the presence of IL-2 triggered vigorous T-cell proliferation and cytokine/growth factor release. These results demonstrate the utility of this skin-explant assay system to further investigate quantitative and qualitative immune responses of NN and PN skin.

scholarly journals Keratinocyte growth factor (KGF) enhances postnatal T-cell development via enhancements in proliferation and function of thymic epithelial cells

Blood ◽  
2007 ◽  
Vol 109 (9) ◽  
pp. 3803-3811 ◽  
Author(s):  
Simona W. Rossi ◽  
Lukas T. Jeker ◽  
Tomoo Ueno ◽  
Sachiyo Kuse ◽  
Marcel P. Keller ◽  
...  
Keyword(s):  
T Cell ◽  
Growth Factor ◽  
Epithelial Cells ◽  
Stromal Cells ◽  
Keratinocyte Growth Factor ◽  
T Cell Development ◽  
Cell Development ◽  
Thymic Epithelial Cells ◽  
Thymic Stromal Cells ◽  
Architectural Organization

Abstract The systemic administration of keratinocyte growth factor (KGF) enhances T-cell lymphopoiesis in normal mice and mice that received a bone marrow transplant. KGF exerts protection to thymic stromal cells from cytoablative conditioning and graft-versus-host disease–induced injury. However, little is known regarding KGF's molecular and cellular mechanisms of action on thymic stromal cells. Here, we report that KGF induces in vivo a transient expansion of both mature and immature thymic epithelial cells (TECs) and promotes the differentiation of the latter type of cells. The increased TEC numbers return within 2 weeks to normal values and the microenvironment displays a normal architectural organization. Stromal changes initiate an expansion of immature thymocytes and permit regular T-cell development at an increased rate and for an extended period of time. KGF signaling in TECs activates both the p53 and NF-κB pathways and results in the transcription of several target genes necessary for TEC function and T-cell development, including bone morphogenetic protein 2 (BMP2), BMP4, Wnt5b, and Wnt10b. Signaling via the canonical BMP pathway is critical for the KGF effects. Taken together, these data provide new insights into the mechanism(s) of action of exogenous KGF on TEC function and thymopoiesis.

Prognostic Analysis of Pre-Transplant CD26-Positive Lymphocytes and CD26bright/CD45RO-Positive Memory T-Cell Levels in Patients Receiving an Autologous Hematopoietic Progenitor-Cell Transplant.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1653-1653
Author(s):  
Martin Hildebrandt ◽  
Hella Gollasch ◽  
Kirstin Rautenberg ◽  
Bernd Doerken ◽  
Wolf-Dieter Ludwig
Keyword(s):  
T Cells ◽  
T Cell ◽  
Progenitor Cell ◽  
Memory T Cells ◽  
Cell Subset ◽  
Hematopoietic Progenitor Cell ◽  
Cell Transplant ◽  
Hematopoietic Progenitor ◽  
Memory T Cell ◽  
Kinetics Of

Abstract The lymphocyte surface glycoprotein CD26 is involved in an array of diverse signalling pathways and costimulatory events. CD26positive cells possess dipeptidylpeptidase IV (DPP IV) activity, a serine protease known to inactivate SDF-1/CXCL12, a key mediator of stem cell homing and engraftment. Furthermore, CD26 modulates surface expression of CTLA-4 and contributes to T-cell migration through endothelial layers. CD26 has been attributed with a central role in alloantigen-mediated immune pathways and memory T-cell responses. The purpose of this study was to evaluate the levels of a distinct memory T-cell subset, CD26bright/CD45RO-positive, in autologous hematopoietic progenitor-cell transplant (HPCT) recipients. Between 2003 and 2006 we enrolled 42 patients scheduled to undergo high-dose chemotherapy and autologous HPCT (multiple myeloma, n=31; Hodgkin’s Disease, n=3; NHL, n=6; PNET, n=1; AML, n=1). Levels of memory and naïve CD26, CD34, CD4, CD8, as well as co-expression of CD4 or CD8 among CD26bright/CD45RO-positive cells were analyzed before autologous HPCT. In addition, the number of memory and naïve CD26-positive T cells transfused per kg body weight in the progenitor cell harvest were determined. The subsets of CD26-positive cells were correlated with kinetics of engraftment and with the occurrence of disease progression or relapse after autologous transplantation. With regard to kinetics of engraftment, only the number of CD34-positive cells transfused was associated with rapid engraftment (P=0.001). However, CD26-positive cells were of predicitve value for the occurrence of disease progression or relapse. Pre-transplant CD26-positive T-cell levels correlated with progression-free survival (PFS) (P=0.022). Specifically, the number of CD26bright/CD45RO-positive memory T-cells in the autograft correlated with PFS and, in regression analyses, emerged as the only variable predictive for the occurrence of diease progression or relapse (P=0.006). The prognostic effects of pre-transplant CD26bright/CD45RO-positive memory T-cells were independent of the type of disease and of the conditioning regimen applied. The analysis of antigens CD4 and CD8, respecively, on CD26bright/CD45RO-positive T-cells yielded no additional information. Our results suggest that pre-transplant levels of CD26-positive T cells, specifically a memory cell subset of CD26bright cells coexpressing CD45RO, may yield information with regard to outcome in autologous HPCT recipients. These observations may contribute to a prospective identification of those patients at higher risk of relapse, based on their immune status.

scholarly journals Kinetics of immune cell reconstitution predict survival in allogeneic bone marrow and G-CSF–mobilized stem cell transplantation

2019 ◽  
Vol 3 (15) ◽  
pp. 2250-2263 ◽  
Author(s):  
Edmund K. Waller ◽  
Brent R. Logan ◽  
Mingwei Fei ◽  
Stephanie J. Lee ◽  
Dennis Confer ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Immune Reconstitution ◽  
Acute Gvhd ◽  
Immune Cell ◽  
Blood Samples ◽  
T Cell Reconstitution

Abstract The clinical utility of monitoring immune reconstitution after allotransplant was evaluated using data from Blood and Marrow Transplant Clinical Trials Network BMT CTN 0201 (NCT00075816), a multicenter randomized study of unrelated donor bone marrow (BM) vs granulocyte colony-stimulating factor (G-CSF)–mobilized blood stem cell (G-PB) grafts. Among 410 patients with posttransplant flow cytometry measurements of immune cell subsets, recipients of G-PB grafts had faster T-cell reconstitution than BM recipients, including more naive CD4+ T cells and T-cell receptor excision circle–positive CD4+ and CD8+ T cells at 3 months, consistent with better thymic function. Faster reconstitution of CD4+ T cells and naive CD4+ T cells at 1 month and CD8+ T cells at 3 months predicted more chronic graft-versus-host disease (GVHD) but better survival in G-PB recipients, but consistent associations of T-cell amounts with GVHD or survival were not seen in BM recipients. In contrast, a higher number of classical dendritic cells (cDCs) in blood samples at 3 months predicted better survival in BM recipients. Functional T-cell immunity measured in vitro by cytokine secretion in response to stimulation with cytomegalovirus peptides was similar when comparing blood samples from BM and G-PB recipients, but the degree to which acute GVHD suppressed immune reconstitution varied according to graft source. BM, but not G-PB, recipients with a history of grades 2-4 acute GVHD had lower numbers of B cells, plasmacytoid dendritic cells, and cDCs at 3 months. Thus, early measurements of T-cell reconstitution are predictive cellular biomarkers for long-term survival and response to GVHD therapy in G-PB recipients, whereas more robust DC reconstitution predicted better survival in BM recipients.

Hepatocyte growth factor preserves graft-versus-leukemia effect and T-cell reconstitution after marrow transplantation

Blood ◽  
2004 ◽  
Vol 104 (5) ◽  
pp. 1542-1549 ◽  
Author(s):  
Takehito Imado ◽  
Tsuyoshi Iwasaki ◽  
Yasuro Kataoka ◽  
Takanori Kuroiwa ◽  
Hiroshi Hara ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Growth Factor ◽  
Acute Gvhd ◽  
Gene Transfection ◽  
T Cell Reconstitution ◽  
Graft Versus Leukemia ◽  
Hepatocyte Growth ◽  
Graft Versus Leukemia Effect

Abstract Graft-versus-host disease (GVHD) is a major complication of allogeneic bone marrow transplantation (BMT). When GVHD is controlled by T-cell–depleted grafts or immunosuppressants, BM transplant recipients often suffer from an increased rate of leukemic relapse and impaired reconstitution of immunity. Using a mouse BMT model, we investigated the effects of hepatocyte growth factor (HGF) gene transfection on the severity of GVHD, the graft-versus-leukemia effect, and the reconstitution of T cells after BMT. After HGF gene transfer, acute GVHD was reduced, while mature donor T-cell responses to host antigens were preserved, resulting in a significant improvement of leukemia-free survival. HGF gene transfer promoted regeneration of bone marrow–derived T cells and the responsiveness of these cells to alloantigens. Furthermore, HGF preserved the thymocyte phenotype and thymic stromal architecture in mice with GVHD. This suggested that HGF exerts a potent protective effect on the thymus, which in turn promotes reconstitution of bone marrow–derived T cells after allogeneic BMT. These results indicate that HGF gene transfection can reduce acute GVHD preserving the graftversus-leukemia effect, while promoting thymic-dependent T-cell reconstitution after allogeneic BMT.

scholarly journals Engineering Regenerative Thymic Tissues to Restore Long-Term T Cell Lymphopoiesis

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5092-5092
Author(s):  
Hui Gai ◽  
Rafa Gras~Pena ◽  
Yogendra Verma ◽  
Vincent Fateh ◽  
Kazuya Ikeda ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Stromal Cells ◽  
Pluripotent Stem Cells ◽  
Immune Reconstitution ◽  
Thymic Epithelial Cells ◽  
Tcr Repertoire ◽  
Thymic Stromal Cells

Abstract The thymus is a primary lymphoid organ that plays a critical role in the development of adaptive T cell immunity and central tolerance. Bone marrow-derived lymphoid progenitor cells migrate into the thymus and interact with thymic epithelial cells (TECs) through sequential positive and negative selection to mature. Thymus-educated mature T cells express a diverse, MHC-restricted and self-tolerant T cell receptor (TCR) repertoire that protects against infection and prevents autoimmunity. Patients born with congenital thymic aplasia, due to 22q11 Deletion Syndrome, or mutations in TBX1, FOXN1 or CHD7, present with complete absence of T cells and a severe combined immunodeficiency (SCID)-like phenotype. Bone marrow transplantation does not cure the thymic defect in these patients and severe infections occur within the first year of life if left untreated. Allogenic thymus transplantation has provided proof of principle that HLA-unmatched pediatric donor thymic tissues can lead to successful immune reconstitution with the emergence of a diverse TCR V-beta repertoire. However, post-transplant organ-specific autoimmunity remains a major concern. Currently allogeneic thymus transplantation is no longer available in the US leaving a deadly therapeutic void for patients born without thymus. Patient-specific or histocompatible thymic tissues derived from pluripotent stem cells could address the critically unmet need, and also a broader range of clinical applications including immune reconstitution post hematopoietic stem cell transplantation (HSCT) and tolerance induction for solid donor organs. The thymus contains two major non-hematological components: the thymic stromal cells and the extracellular matrix (ECM). The thymic stromal layer is composed of thymic epithelial cells and mesenchymal cells. The thymic ECM forms a three-dimensional (3D) network to provide physical support and nutrition to thymic stromal cells. Methods: To address the need for histocompatible regenerative thymic tissues, we aim to differentiate fully functional thymic epithelial progenitor cells (TEPCs) from human pluripotent stem cells (hPSCs) and further generate 3D transplantable organoids using engineered matrix proteins that mimic the native thymic microenvironment. Results: We have developed a novel platform to generate hPSC-derived TEPCs by dissecting the key signaling pathways that govern human thymic ontogeny. These hPSC-derived TEPCs express the defining markers of TEPC-fate, such as FoxN1, Cytokeratin 8, Cytokeratin 5, Delta-like Canonical Notch Ligand 4 (DLL4) and MHC class II. Previous studies have shown FoxN1 to be the master regulator controlling thymic development, however, little is known about its regulatory network. Elucidating and validating the factors that initiate and maintain FoxN1 expression is the key to successfully engineer sustainable thymic tissues. We have identified a combination of morphogens that can maintain the expression of FoxN1, DLL4 and AIRE of primary TECs in culture. To gain insight into the composition of primary thymic ECM proteins and adapt their characteristics beyond the features of commercially available 3D hydrogels, we analyzed a series of human fetal thymic tissues using whole transcriptome analysis. Our current work focuses on adapting our 2D culture protocol to sustain hPSC-TEPCs in 3D matrix-based organoids. Ongoing studies test the capacity of hPSC-TECPs to promote T cell maturation and the development of a diverse TCR repertoire in an athymic xenograft mouse model (NSG-FoxN1null). Conclusions: hPSC can be differentiated in vitro into TEPC-fate and developed into thymic organoids using custom-designed protein matrices. Studies to test sustainability and functionality of the engineered thymic organoids in vivo are currently under way. Disclosures No relevant conflicts of interest to declare.

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