Abstract A38: The Wnt noncanonical receptor ROR1 regulates neuroblastoma cell growth and motility through two distinct mechanisms

Abstract

Abstract Neuroblastoma is a pediatric tumor derived from the sympathetic nervous system, responsible for about 15% of cancer deaths in children. The Wnt signaling pathway, which is required for both normal embryonic development and the maintenance of different types of adult stem cells, including sympathoadrenal progenitors, plays a major role in the onset and progression of several types of human cancer. In a screen of several human neuroblastoma cell lines, we found that none showed a constitutive activation of the canonical Wnt/β-catenin signaling, arguing against a major role of this pathway in this type of cancer. In a previous study, we demonstrated that the receptor tyrosine kinase-like orphan receptor 1 (ROR1) can be triggered by noncanonical Wnt ligands, resulting in its phosphorylation on serine/threonine residues. We found that several neuroblastoma cells express high levels of ROR1, which can be associated in some cases with constitutive phosphorylation of this receptor through an autocrine mechanism. We showed that ROR1 down-regulation through lentiviral shRNAs strongly reduced the growth of different neuroblastoma cells, including SJNB-10, SK-N-AS and Kelly. Conversely, ROR1 knockdown in other neuroblastoma cells significantly reduced cell motility, without affecting growth. To assess the potential role of autocrine noncanonical Wnt signaling in neuroblastoma cell motility and/or growth, we used two different approaches, based on the inhibition of Wnt secretion using the small molecule IWP2, or the sequestration of Wnt ligands by ext-Fzd8, a decoy receptor containing the extracellular domain of the Wnt co-receptor Frizzled 8. These studies revealed that IWP-2 and ext-Fzd8 could both inhibit ROR1 constitutive phosphorylation and reduce cell motility, but they had no effect on cell growth, indicating that different mechanisms are responsible for the activation of this receptor in neuroblastoma cells. It has been recently reported that the putative tyrosine kinase domain of ROR1 could play a role in the survival of lung cancer cells. We found that the inactivation of ROR1 ATP-binding domain using the CRISPR/Cas9 technology did not affect the growth of SJNB-10 and Kelly cells, implying that the catalytic activity of this receptor is not required for its effects in neuroblastoma cells. In conclusion, we demonstrated that ROR1 can play a dual role in neuroblastoma cell motility and growth through two distinct mechanisms, respectively dependent and independent of noncanonical Wnt ligands, suggesting that this receptor could represent a novel therapeutic target for the treatment of this type of cancer. Citation Format: Alexis Guernet, Sardar Faisal Mahmood, Hassan Ainani, Youssef Anouar, Luca Grumolato. The Wnt noncanonical receptor ROR1 regulates neuroblastoma cell growth and motility through two distinct mechanisms. [abstract]. In: Proceedings of the Fourth AACR International Conference on Frontiers in Basic Cancer Research; 2015 Oct 23-26; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2016;76(3 Suppl):Abstract nr A38.