Abstract 3266: Inhibition of Rho-associated kinase 2 suppresses MYCN expression and induces differentiation of neuroblastoma
Authors: Cecilia Dyberg, David Forsberg, Susanne Fransson, Jessika Lännerholm Palm, Bjorn Helge Haug, Baldur Sveinbjornsson, Tommy Martinsson, Per Kogner, John Inge Johnsen, Malin Wickstrom
Journal: Cancer Research (2015)

Abstract Background: Neuroblastoma is an embryonic tumor of the peripheral sympathetic nervous system that originates from cells within the neural crest. The non-canonical Wnt/planar cell polarity (PCP) signaling pathway regulates cytoskeletal organization, migration and maturation of neural crest cells during neuritogenesis. We and others have shown that high-risk neuroblastoma exhibit frequent mutations of genes controlling the activity of the Wnt/PCP signaling cascade leading to inhibition of Rac and activation of Rho resulting in downstream activation of the serine/threonine kinases Rho-Associated Coiled-Coil Containing Protein Kinases (ROCK1 and ROCK2) Methods: We performed whole-exome sequencing for 33 neuroblastoma samples of all clinical subgroups to search for mutations and genetic aberrations. Cytotoxic activity of ROCK inhibitors was studied in cell viability assays. Morphology and invasion were studied with microscopy. The molecular mechanisms were characterized using cell- and molecular biology techniques. In vivo studies (xenografts and the transgenic mouse model TH-MYCN) in mice were carried out to validate the therapeutic effects and toxicity. Results: Exome sequencing detected frequent mutations and gene aberrations in genes controlling the activity of PCP signalling. Analysis ofgene signatures and immunohistochemistry showed that high expression of ROCK1 and ROCK2 correlated with poor patient survival. Several mediators of Rho/Rac were differentially expressed in cell lines and patient samples. Using compounds blocking ROCK1 and ROCK2 activity revealed that the ROCK2 inhibitor HA-1077 effectively repressed proliferation and reduced cell viability in neuroblastoma. HA-1077 inhibited migration and induced differentiation through inhibition of neural outgrowth and inhibited MYCN protein expression, effects that were mimicked using siRNA against ROCK. Finally, HA-1077 reduced the growth of established neuroblastoma xenografts in nude mice and repressed tumor progression in a MYCN-driven mouse model of neuroblastoma (T-MYCN) Conclusions: Our results show the significance of the non-canonical/PCP signaling cascade in neuroblastoma development and progression. Furthermore, ROCK inhibition suppress the growth of neuroblastoma in preclinical models and provide a novel option for improved treatment of high-risk neuroblastoma. Citation Format: Cecilia Dyberg, David Forsberg, Susanne Fransson, Jessika Lännerholm Palm, Bjorn Helge Haug, Baldur Sveinbjornsson, Tommy Martinsson, Per Kogner, John Inge Johnsen, Malin Wickstrom. Inhibition of Rho-associated kinase 2 suppresses MYCN expression and induces differentiation of neuroblastoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3266. doi:10.1158/1538-7445.AM2015-3266