About

Paul J Gadue, PhD, is a Professor of Pathology and Laboratory Medicine at the Perelman School of Medicine at the University of Pennsylvania. He serves as Associate Director of the Human Pluripotent Stem Cell Core Facility at Children's Hospital of Philadelphia and is a member of several research centers including the Center for Cellular and Molecular Therapeutics, the Institute for Regenerative Medicine, and the Diabetes and Endocrinology Research Center. His research focuses on human development, specifically endoderm and mesoderm specification, utilizing human embryonic stem (ES) cells and induced pluripotent stem (iPS) cells. His laboratory studies cell fate decisions during embryogenesis, aiming to understand the molecular mechanisms regulating endoderm and mesoderm development through in vitro differentiation models. A key area of his work involves investigating hematopoiesis with an emphasis on megakaryocyte development, with the goal of optimizing platelet generation from ES/iPS cells and developing disease models for platelet disorders using patient-derived iPS cells and CRISPR/Cas9 technology. Additionally, his research explores endoderm formation, pancreatic, liver, and lung specification, and the transcriptional control of these processes, with a particular focus on generating functional pancreatic beta cells and modeling genetic forms of diabetes.

Research topics

• Biology
• Cell biology
• Genetics
• Immunology
• Cancer research

Selected publications

• Generation of CHOPi014-A from healthy adult peripheral blood mononuclear cells

  Stem Cell Research · 2025-08-24

  articleOpen access

  CHOPWT15is a control male induced pluripotent stem cell (iPSC) line that can be used to model genetic variants by creating an allelic series of isogenic lines using genome editing technologies (Hockemeyer and Jaenisch, 2016; Maguire et al., 2022; Pavani et al., 2022). An allelic series of iPSC lines provides the means to perform genotype and phenotype analyses of pathological variants without the confounding effects of genetic background. Peripheral blood mononuclear cells (PBMCs), obtained from a healthy adult male, were reprogrammed using Sendai virus creating an iPSC line that has undetectable copy number variations (CNVs, resolution >= 100 kb) and differentiates to all three germlayers.

  PublisherDOI

• Using iPSC-derived hematopoietic stem cells with long-term engraftment capability to model fetal blood disorders

  Blood · 2025-11-03

  articleOpen access

  Abstract Introduction: Human developmental hematopoiesis is a complex process occurring in sequential waves at different embryonic sites. This process yields both differentiated blood cells essential for embryonic development and hematopoietic stem cells (HSCs) necessary for lifelong blood cell production. There are at least two main hematopoietic programs during embryogenesis: a transient primitive wave that primarily generates myelo-erythroid progenitors in the yolk sac, and a definitive wave occurring in the aorta-gonad-mesonephros region which produces progenitors with expanded lineage potential and the first transplantable HSCs. A challenge in generating HSCs from induced pluripotent stem cells (iPSCs) is that many current methods recapitulate early developmental stages only, failing to produce transplantable HSCs and limiting their application for in vivo studies. We have previously shown that we can recapitulate fetal erythropoiesis using a protocol that produces definitive hematopoietic cells. Red cells generated with this method primarily produce fetal globin and are functionally distinct from primitive erythroblasts. Here, we adapted this method to a serum-free 3D culture system to produce definitive hematopoietic stem progenitor cells (HSPCs) from iPSCs to 1) evaluate long-term engraftment via xenotransplantation and 2) model a preleukemic disorder of fetal origin, transient abnormal myelopoiesis (TAM), which affects ~20% of neonates with Trisomy 21 (T21) and is associated with mutations in the key hematopoietic transcription factor GATA1. Methods We designed a serum-free 3D culture system that directs mesodermal commitment through Wnt pathway activation by manipulating developmental cues through retinoic acid signaling and shear stress. We transplanted 24 NBSGW mice with 1.2-2 million iPSC-derived HSPCs using 3 distinct wildtype (WT) lines and experimental batches. The 3D definitive differentiation was assessed using isogenic T21 iPSCs with wild type GATA1 or the truncated isoform lacking the N-terminus, GATA1s. Results Long-term repopulation was achieved up to 20 weeks post-transplant in the bone marrow (BM) of all engrafted mice (mean = 3.9±11.7% human HLA-ABC+) and in the peripheral blood of 16/24 mice. In mice with the highest level of engraftment (>10%), we detected human HSCs as well as myeloid, lymphoid, erythroid and megakaryocyte precursors in the BM, indicating complete hematopoietic reconstitution. Human CD34+ cells recovered from the graft retained multilineage potential in colony forming assays. Importantly, no leukemia or teratomas were observed in any of the transplanted mice. To assess whether this definitive iPSC culture protocol could faithfully model the fetal blood disorder TAM, we generated definitive HSPCs from T21/wtGATA1 and T21/GATA1s iPSCs. Both T21 lines generated budding HSPCs at day 15; however, the percentage of CD34+CD45+ HSPCs was lower compared to euploid controls (60.5% of WT ±12.9 for T21, and 64.5% of WT ± 0.06 for T21/GATA1s). In addition, at day 15 we observed a significant CD41+CD42b+ megakaryocyte population in both trisomy lines, consistent with the disease phenotype and that was not previously observed with our primitive hematopoietic differentiation protocols. Compared to WT controls, we observed a 2.3-fold and a 4.2-fold increased megakaryocyte population from T21/wtGATA1 and T21/GATA1s iPSCs, respectively. T21/GATA1s HPSCs also showed aberrant morphology with megakaryocytic and blast-like features. Conclusions These studies show that our serum-free 3D culture system can generate iPSC-derived HSPCs that can engraft immunodeficient mice, reconstitute different blood lineages, and persist for 20 weeks. Using a T21/TAM iPSC model, we found an enhanced megakaryocyte population in T21/wtGATA1 consistent with a phenotype we observed with primary human fetal liver hematopoiesis, but not with iPSC-derived primitive hematopoietic progenitors. Thus, T21/GATA1s definitive HSPCs recapitulate the enhanced megakaryopoiesis consistent with TAM in vitro. Ongoing xenotransplant experiments will help elucidate the interaction of trisomy 21 and GATA1s and provide a novel in vivo model to study and treat human hematological diseases.

  PublisherOA PDFDOI

• 3048 – NOD1-DEPENDENT NF-KB ACTIVATION INITIATES HEMATOPOIETIC STEM CELL SPECIFICATION IN RESPONSE TO SMALL RHO GTPASES

  Experimental Hematology · 2024-08-01

  article
  PublisherDOI

• Tropomyosin 1 deficiency facilitates cell state transitions and enhances hemogenic endothelial cell specification during hematopoiesis

  Stem Cell Reports · 2024-08-29 · 6 citations

  articleOpen access

  Tropomyosins coat actin filaments to impact actin-related signaling and cell morphogenesis. Genome-wide association studies have linked Tropomyosin 1 (TPM1) with human blood trait variation. TPM1 has been shown to regulate blood cell formation in vitro, but it remains unclear how or when TPM1 affects hematopoiesis. Using gene-edited induced pluripotent stem cell (iPSC) model systems, we found that TPM1 knockout augmented developmental cell state transitions and key signaling pathways, including tumor necrosis factor alpha (TNF-α) signaling, to promote hemogenic endothelial (HE) cell specification and hematopoietic progenitor cell (HPC) production. Single-cell analyses revealed decreased TPM1 expression during human HE specification, suggesting that TPM1 regulated in vivo hematopoiesis via similar mechanisms. Analyses of a TPM1 gene trap mouse model showed that TPM1 deficiency enhanced HE formation during embryogenesis, without increasing the number of hematopoietic stem cells. These findings illuminate novel effects of TPM1 on developmental hematopoiesis.

  PublisherDOI

• Generation of a fluorescent mNeonGreen insulin reporter line in the H1 (WA01) hESC background

  Stem Cell Research · 2024-09-11 · 1 citations

  articleOpen accessSenior author

  Over the past decade, the use of human stem cell-derived β cells (SC-β cells) to model pancreatic β cell development, function and disease has become increasingly common. Though protocols are rapidly improving, current directed differentiation strategies do not yield a pure population of insulin-positive SC-β cells in vitro. Therefore, it is experimentally advantageous to have reporter lines that allow for live sorting of insulin-positive populations. To aid in these studies, we have knocked mNeonGreen fluorescent protein into the endogenous insulin locus of the commonly used H1 (WA01) human embryonic stem cell line.

  PublisherDOI

• Modeling primitive and definitive erythropoiesis with induced pluripotent stem cells

  Blood Advances · 2024-01-30 · 15 citations

  articleOpen accessSenior author

  ABSTRACT: During development, erythroid cells are produced through at least 2 distinct hematopoietic waves (primitive and definitive), generating erythroblasts with different functional characteristics. Human induced pluripotent stem cells (iPSCs) can be used as a model platform to study the development of red blood cells (RBCs) with many of the differentiation protocols after the primitive wave of hematopoiesis. Recent advances have established that definitive hematopoietic progenitors can be generated from iPSCs, creating a unique situation for comparing primitive and definitive erythrocytes derived from cell sources of identical genetic background. We generated iPSCs from healthy fetal liver (FL) cells and produced isogenic primitive or definitive RBCs which were compared directly to the FL-derived RBCs. Functional assays confirmed differences between the 2 programs, with primitive RBCs showing a reduced proliferation potential, larger cell size, lack of Duffy RBC antigen expression, and higher expression of embryonic globins. Transcriptome profiling by scRNA-seq demonstrated high similarity between FL- and iPSC-derived definitive RBCs along with very different gene expression and regulatory network patterns for primitive RBCs. In addition, iPSC lines harboring a known pathogenic mutation in the erythroid master regulator KLF1 demonstrated phenotypic changes specific to definitive RBCs. Our studies provide new insights into differences between primitive and definitive erythropoiesis and highlight the importance of ontology when using iPSCs to model genetic hematologic diseases. Beyond disease modeling, the similarity between FL- and iPSC-derived definitive RBCs expands potential applications of definitive RBCs for diagnostic and transfusion products.

  PublisherOA PDFDOI

• p65 signaling dynamics drive the developmental progression of hematopoietic stem and progenitor cells through cell cycle regulation

  Nature Communications · 2024-09-06 · 11 citations

  articleOpen access

  Most gene functions have been discovered through phenotypic observations under loss of function experiments that lack temporal control. However, cell signaling relies on limited transcriptional effectors, having to be re-used temporally and spatially within the organism. Despite that, the dynamic nature of signaling pathways have been overlooked due to the difficulty on their assessment, resulting in important bottlenecks. Here, we have utilized the rapid and synchronized developmental transitions occurring within the zebrafish embryo, in conjunction with custom NF-kB reporter embryos driving destabilized fluorophores that report signaling dynamics in real time. We reveal that NF-kB signaling works as a clock that controls the developmental progression of hematopoietic stem and progenitor cells (HSPCs) by two p65 activity waves that inhibit cell cycle. Temporal disruption of each wave results in contrasting phenotypic outcomes: loss of HSPCs due to impaired specification versus proliferative expansion and failure to delaminate from their niche. We also show functional conservation during human hematopoietic development using iPSC models. Our work identifies p65 as a previously unrecognized contributor to cell cycle regulation, revealing why and when pro-inflammatory signaling is required during HSPC development. It highlights the importance of considering and leveraging cell signaling as a temporally dynamic entity.

  PublisherOA PDFDOI

• Tropomyosin 1 deficiency facilitates cell state transitions to enhance hemogenic endothelial cell specification during hematopoiesis

  bioRxiv (Cold Spring Harbor Laboratory) · 2023-09-02

  preprintOpen access

  Abstract Tropomyosins coat actin filaments and impact actin-related signaling and cell morphogenesis. Genome-wide association studies have linked Tropomyosin 1 ( TPM1 ) with human blood trait variation. Prior work suggested that TPM1 regulated blood cell formation in vitro, but it was unclear how or when TPM1 affected hematopoiesis. Using gene-edited induced pluripotent stem cell (iPSC) model systems, TPM1 knockout was found to augment developmental cell state transitions, as well as TNFα and GTPase signaling pathways, to promote hemogenic endothelial (HE) cell specification and hematopoietic progenitor cell (HPC) production. Single-cell analyses showed decreased TPM1 expression during human HE specification, suggesting that TPM1 regulated in vivo hematopoiesis via similar mechanisms. Indeed, analyses of a TPM1 gene trap mouse model showed that TPM1 deficiency enhanced the formation of HE during embryogenesis. These findings illuminate novel effects of TPM1 on developmental hematopoiesis.

  PublisherOA PDFDOI

• Synergistic roles of DYRK1A and GATA1 in trisomy 21 megakaryopoiesis

  JCI Insight · 2023-10-31 · 13 citations

  articleOpen access

  Patients with Down syndrome (DS), or trisomy 21 (T21), are at increased risk of transient abnormal myelopoiesis (TAM) and acute megakaryoblastic leukemia (ML-DS). Both TAM and ML-DS require prenatal somatic mutations in GATA1, resulting in the truncated isoform GATA1s. The mechanism by which individual chromosome 21 (HSA21) genes synergize with GATA1s for leukemic transformation is challenging to study, in part due to limited human cell models with wild-type GATA1 (wtGATA1) or GATA1s. HSA21-encoded DYRK1A is overexpressed in ML-DS and may be a therapeutic target. To determine how DYRK1A influences hematopoiesis in concert with GATA1s, we used gene editing to disrupt all 3 alleles of DYRK1A in isogenic T21 induced pluripotent stem cells (iPSCs) with and without the GATA1s mutation. Unexpectedly, hematopoietic differentiation revealed that DYRK1A loss combined with GATA1s leads to increased megakaryocyte proliferation and decreased maturation. This proliferative phenotype was associated with upregulation of D-type cyclins and hyperphosphorylation of Rb to allow E2F release and derepression of its downstream targets. Notably, DYRK1A loss had no effect in T21 iPSCs or megakaryocytes with wtGATA1. These surprising results suggest that DYRK1A and GATA1 may synergistically restrain megakaryocyte proliferation in T21 and that DYRK1A inhibition may not be a therapeutic option for GATA1s-associated leukemias.

  PublisherOA PDFDOI

• Generation of a human <i>Tropomyosin 1</i> knockout iPSC line

  bioRxiv (Cold Spring Harbor Laboratory) · 2023-05-04 · 1 citations

  preprintOpen access

  Abstract The CHOPWT17_TPM1KOc28 iPSC line was generated to interrogate the functions of Tropomyosin 1 ( TPM1 ) in primary human cell development. This line was reprogrammed from a previously published wild type control iPSC line.

  PublisherOA PDFDOI

Recent grants

• HNF1A in human endocrine cell development and function

  NIH · $2.1M · 2020–2025

• Microphysiological systems for modeling autoimmunity in type 1 diabetes

  NIH · $2.3M · 2019–2024

• NIH Grant F32HL076058

  NIH · $140k · 2006

• NIH Grant UC4DK104196

  NIH · $72k · 2019

• NIH Grant R01HL130698

  NIH · $2.2M · 2020

Frequent coauthors

• Deborah L. French

  174 shared

• Jean Ann Maguire

  Children's Hospital of Philadelphia

  65 shared

• Alyssa Gagne

  50 shared

• Jason A. Mills

  University of California, Los Angeles

  37 shared

• Fabian L. Cardenas‐Diaz

  University of Pennsylvania

  35 shared

• Mortimer Poncz

  Children's Hospital of Philadelphia

  32 shared

• Sara Borst

  University of Pennsylvania

  28 shared

• Stella T. Chou

  University of Pennsylvania

  27 shared

Labs

• Pathology and Laboratory Medicine, University of PennsylvaniaPI