Retinoic acid induces differentiation in neuroblastoma via ROR1 by modulating retinoic acid response elements

Abstract

Neuroblastoma is the most common inheritable, solid neoplasm in children found under the age of 7 and accounts for approximately 7% of childhood cancers. A common treatment that has been prescribed for over a decade is retinoid therapy [using all-trans retinoic acid (RA)]. Treatment with this differentiating agent has been revealed to progress the cells from their stem-cell state to a mature neuronal state gaining classical neuronal characteristics, including the suppression of proliferation. However, the molecular mechanism underlying the action of RA treatment remains to be elucidated. In the present study, a novel mechanism of RA-induced differentiation via regulation of receptor tyrosine kinase-like orphan receptor 1 (ROR1) is reported. ROR1 is overexpressed in neuroblastoma but significantly downregulated in mature differentiated neurons. Hence, it was hypothesized that RA may modulate ROR1 leading to differentiation and termination of cancerous properties. Immunoblotting revealed that following RA treatment, ROR1 levels initially increased then sharply decreased by 96 h. This was paired with synaptophysin, a mature neuron marker, sharply increasing concurrently, providing evidence of differentiation by 96 h. Investigation of the ROR1 pathway confirmed ROR1-dependent downstream activation of the PI3K/AKT signaling axis, a growth pathway previously demonstrated to promote differentiation. Chromatin immunoprecipitation revealed an increase in RAR binding to the promoters of ROR1 and its endogenous ligand, Wnt5a. This research provided compelling evidence that RA is able to modulate the expression of ROR1 and Wnt5a to promote differentiation through the expression of synaptophysin. This data combined with the overarching data from the scientific community regarding proliferation and other proliferative factors in early-stage neurons provides a more in-depth model of the process of differentiation in neurons.

Systematic review of the receptor tyrosine kinase superfamily in neuroblastoma pathophysiology

“…Altogether, we found that RYK and ROR2 , but not ROR1 , are robustly associated with worse outcome in neuroblastoma. This is further supported by a recent study describing a role for ROR1, but not ROR2, in retinoic acid-induced neuroblastoma differentiation [ 145 ]. Together, these data suggest that the WNT modifying pseudokinases ROR2 and RYK represent potential novel therapeutic targets for neuroblastoma treatment and that current research on immuno-therapeutic strategies against ROR1 might need to be re-evaluated in favor of ROR2 and RYK.…”

Restoration of functional PAX3 transcriptional factor enhanced neuronal differentiation in PAX3b isoform-depleted neuroblastoma cells

“…Similarly, the earlier research work also showed no induction of neuronal phenotype upon RA treatment in SK-N-SH cells rather it converts these cells into epithelial-like morphology (Sidell et al 1983). In contrast to these observations, the recent article claimed that RA treatment induced neuronal differentiation in SK-N-SH cells as shown by neuronal marker SYP expression (Illendula et al 2020). However, their data showed more than 50% reduction of SYP in 48 hours of differentiation which is in contrast to their claim (Illendula et al 2020) and we have reported a similar reduction of TUBB3 by RA treatment in SK-N-SH cells (Figure 2F).…”

“…In contrast to these observations, the recent article claimed that RA treatment induced neuronal differentiation in SK-N-SH cells as shown by neuronal marker SYP expression (Illendula et al 2020). However, their data showed more than 50% reduction of SYP in 48 hours of differentiation which is in contrast to their claim (Illendula et al 2020) and we have reported a similar reduction of TUBB3 by RA treatment in SK-N-SH cells (Figure 2F). Further, another study by Liang Y and co-workers also could not nd RA-mediated induction of TUBB3 or SYP in SK-N-SH cells (Liang et al 2009).…”

NME1 Protects Against Neurotoxin-, α-Synuclein- and LRRK2-Induced Neurite Degeneration in Cell Models of Parkinson’s Disease

“…NME1 has been reported to interact with RORa [33], and RORa is thought to be a candidate gene for sporadic PD [53]. Additionally, as retinoic acid exerts its differentiationstimulating effects through ROR2 and RORa in cultured mouse hippocampal neurons and neuroblastoma cell lines [54][55][56], we examined the roles of both RORa and ROR2 receptors in NME1-promoted neurite growth. We found that NME1 required both RORa and ROR2 to promote neurite growth in SH-SY5Y cells.…”