Abstract
Neuroblastoma is a cancer of neural crest stem cell (NCSC) lineage. Signaling pathways that regulate NCSC differentiation have been implicated in neuroblastoma tumorigenesis. This is exemplified by MYCN oncogene targets that balance proliferation, differentiation, and cell death similarly in normal NCSC and in high-risk neuroblastoma. Our previous work discovered a survival mechanism by which MYCN-amplified neuroblastoma circumvents cell death by upregulating components of the error-prone non-canonical alternative nonhomologous end-joining (alt-NHEJ) DNA repair pathway. Similar to proliferating stem cells, high-risk neuroblastoma cells have enhanced DNA repair capacity, overcoming DNA damage with higher repair efficiency than somatic cells. Adequate DNA maintenance is required for lineage protection as stem cells proliferate and during tumor progression to overcome oncogene-induced replication stress. On this basis, we hypothesized that alt-NHEJ overexpression in neuroblastoma is a cancer cell survival mechanism that originates from DNA repair systems of NCSC, the presumed progenitor cell of origin. A human NCSC model was generated in which inducible MYCN triggered an immortalized phenotype capable of forming metastatic neuroectodermal tumors in mice, resembling human neuroblastoma. Critical alt-NHEJ components (DNA Ligase III, DNA Ligase I, and Poly [ADP-ribose polymerase 1]) were highly expressed in normal early NCSC, and decreased as cells became terminally differentiated. Constitutive MYCN expression maintained high alt-NHEJ protein expression, preserving the expression pattern of the immature neural phenotype. siRNA knockdown of alt-NHEJ components reversed MYCN effects on NCSC proliferation, invasion, and migration. DNA Ligase III, Ligase I, and PARP1 silencing significantly decreased neuroblastoma markers expression (TH, Phox2b, and TRKB). These results utilized the first human NCSC model of neuroblastoma to uncover an important link between MYCN and alt-NHEJ expression in developmental tumor initiation, setting precedence to investigate alt-NHEJ repair mechanics in neuroblastoma DNA maintenance.
Highlights
Neuroblastoma (NBL), the most common extracranial tumor in children, is thought to arise from neural crest progenitor cells[1]
neural crest stem cell (NCSC) isolated by this method have been previously characterized as multipotent and capable of differentiation into neural crest derivatives including neurons, Schwann cells, myofibroblasts, and sympathoadrenal cells when exposed to differentiation media. human embryonic stem cells (hESC)-derived cells were FACSsorted after 8 days to collect cells that stained double positive for NCSC markers p75 and HNK-1 (Fig. 1b)
This study provides the first hESC model of NBL derived from transformation of normally differentiating human NCSC, and further implicates neural crest progenitors as the cellof-origin of NBL tumors
Summary
Neuroblastoma (NBL), the most common extracranial tumor in children, is thought to arise from neural crest progenitor cells[1]. Signaling pathways critical for normal neural crest stem cell (NCSC) development have been implicated in NBL pathogenesis, maintaining unique embryonic properties that balance migration, proliferation, differentiation, and cell death[2]. This is highlighted by Official journal of the Cell Death Differentiation Association. In the presence of cellular stress and DNA damaging agents, an attenuated G1 checkpoint suggests that surviving NBL cells require efficient DNA maintenance pathways that circumvent apoptosis and avoid senescence[10] This unique characteristic is distinct from somatic cells, but similar to rapidly proliferating embryonic stem cells that must maintain an effective
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