A hydrogel-based 3D culture model for Down syndrome-myeloid leukemia and ex vivo drug testing
Date
2021
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
University of Delaware
Abstract
Induced pluripotent stem cells (iPSCs) provide an extraordinary tool for disease modeling owing to their potential to differentiate into the desired cell type. The differentiation of iPSCs is typically performed on 2-dimensional monolayers of stromal cell or animal tissue derived extracellular matrices. Recent advancements in disease modeling have utilized iPSCs in 3-dimensional (3D) cultures to study diseases such as muscular dystrophy, cardiomyopathy, pulmonary fibrosis, and wound healing. However, these approaches are yet to be explored in modeling the hematological malignancies such as acute myeloid leukemia in Down syndrome childrem (DS-ML). Genomic characterization of DS-ML blasts showed the presence of a unique mutation in GATA1, an essential hematopoietic transcription factor, leading to the production of a truncated from of GATA1 (GATA1s). GATA1s together with trisomy 21 is sufficient to develop a pre-leukemic condition called transient myeloproliferative disorder (TMD). Approximately thirty percent of these cases progress into DS-ML by acquisition of additional somatic mutations in a stepwise manner. STAG2, an accessory cohesin complex component, is often mutated in DS-ML. In this study, I established a synthetic 3D iPSC culture system for modeling DS-ML via hematopoietic differentiation of customizable iPSCs. A chemically cross-linkable PEG hydrogel decorated with integrin binding peptide was found to be permissive of hematopoietic differentiation of iPSCs. It provided a cost-effective system for the generation of hematopoietic stem and progenitor cells with higher yield of early HSPCs compared to traditional 2D culture on Matrigel coated dishes. Characterization of the cell populations in the HSPCs produced from the iPSC lines harboring mutation in GATA1 and STAG2 cultured in 3D showed that the erythroid population was reduced whereas the megakaryoid and myeloid populations were significantly increased in mutant trisomic line compared to disomic or trisomic lines with wild-type GATA1 and STAG2, consistent with DS-ML characteristics. In conclusion, I have identified a cost-effective tunable 3D hydrogel system to model DS-ML. ☐ While DS-ML has favorable outcome to chemotherapy, it is still plagued by relapse in 20% of patients. The occurrence of this relapse can be attributed to chemoprotection provided by the bone marrow (BM) microenvironment (cellular and extracellular matrix components) and epigenetic alterations accumulated during leukemogenesis. The 2D in vitro drug testing does not recapitulate the cellular interactions in BM mimicking environment. In this study, I established a synthetic 3D ex-vivo model mimicking the extracellular matrix components, mechanics and one of the key cellular components (mesenchymal stem cells, MSCs) of BM microenvironment. The BM-MSCs induce chemoprotection to cytotoxic drugs in DS-ML cells. This chemoprotection can be overridden by use of epigenetic modifier azacitidine (Aza, DNA methytransferase inhibitor). This study provides proof of principle for establishing a BM mimicking ex vivo model and can be utilized for high throughput drug screening.
Description
Keywords
Induced pluripotent stem cells (iPSCs), Disease modeling