Developing new models of childhood malignancies using human induced pluripotent stem cells

Sammanfattning: Early onset diseases such as childhood malignancies and neurodevelopmental disorders have been intricate to study. For many years, research has relied and dependent upon different animal systems. Despite the usefulness of these systems, which have allowed the understanding of the biology behind these processes, the differences between species are still an undoubted fact. The emergence of induced pluripotent stem (iPS) cell technology has indeed opened new venues for many fields including disease modeling, personalized cell therapy, and drug screening. iPS cells have the potential to virtually differentiate into any cell type, hence becoming an unlimited source of disease-relevant cell types. Here, we present examples demonstrating the potential of disease modeling using patient-derived iPS cells. Neuroblastoma (NB) and Medulloblastoma (MB) are both cancers linked to dysregulations in pathways important during human development. Whereas NB develops during the peripheral nervous system (PNS) development, MB initiates during central nervous system (CNS) development. We have taken advantage of the early developmental signature of iPS cells to model cancer. We used non-cancerous cells from patients carrying germline mutations in cancer predisposing genes, ALK and PTCH1, and developed in vivo models that offer a unique understanding of cancer initiation and progression. NB patients carrying an ALK germline mutation were used to generate iPS cells (Paper I) and subsequent differentiation to Neural Crest Cells (NCC) was performed (Paper II, III). For this, a NCC generation protocol was optimized using intermediate levels of BMP (Paper II). Next, labelled NCC from patients and controls were orthotopically transplantated into the adrenal gland of immunodeficient mice (Paper III). Mice were followed in vivo using IVIS system, and we detected increased luciferase signal after more than 8 weeks but no signal was observed in mice injected with control NCC. After a year, adrenal glands from mice were harvested and one case of ganglioneuroblastoma was diagnosed, suggesting a low penetrance and mild phenotype of ALK contribution in NB initiation. Using a similar workflow, we generated Neuroepithelial stem (NES) cells from iPS cells derived from Gorlin syndrome patients. Gorlin patients carry germline mutations in PTCH1. Mutations in PTCH1 constitutively activate the Sonic Hedgehog (SHH) signalling pathway. In vivo transplantation of patient cells into the cerebellum of immunocompromised mice showed faithful resemblance of human SHH MB. By establishing NES cell cultures derived from MB tumors in the mice cerebellum, we could show the potential use of this model for identifying new targets for cancer treatment (Paper IV).b Moreover, we exploited 2D and 3D human in vitro systems derived from iPS cells to study the role of p53 during early brain development. We show that p53 has an important function in maintaining the appropriate structure of human brain organoids. Moreover, we demonstrate that p53 maintains genomic instability and primes neural differentiation in human NES cells. Thus, revealing the role of p53 in a human in vitro context of brain development (Paper V). In summary our work presents the big potential of iPS cell technology in the field of modeling disease.