Targeting the CALCB/RAMP1 axis inhibits growth of Ewing sarcoma

• DOI: 10.1038/s41419-019-1372-0
• Publication Date: 2019-02-11T09:04:22Z
• Authors: Marlene Dallmayer; Jing Li; Shunya Ohmura; Rebeca Alba Rubio; Michaela C. Baldauf; Tilman L. B. Hölting; Julian Musa; Max M. L. Knott; Stefanie Stein; Florencia Cidre-A...; Fabienne S. Wehweck; Laura Romero-Pérez; Julia S. Gerke; Martin F. Orth; Aruna Marchetto; Thomas Kirchner; Horacio Bach; Giuseppina Sannino; Thomas G. P. Grün...
• Keywords: FLI1

Ewing sarcoma (EwS) is an aggressive cancer characterized by chromosomal translocations generating fusions of the EWSR1 gene with ETS transcription factors (in 85% FLI1). EWSR1-FLI1 induces expression via binding to enhancer-like GGAA-microsatellites, whose activity correlates number consecutive GGAA-repeats. Herein we investigate role secretory neuropeptide CALCB (calcitonin-related polypeptide β) in EwS, which signals CGRP (calcitonin gene-related peptide) receptor complex, containing RAMP1 (receptor modifying protein 1) as crucial part for specificity. Analysis 2678 microarrays comprising 50 tumor entities and 71 normal tissue types revealed that specifically highly overexpressed EwS. Time-course knockdown experiments showed tightly linked EWSR1-FLI1. Consistently, set enrichment analyses genes primary EwS correlated indicated it co-expressed other target associated signatures involved stemness proliferation. Chromatin immunoprecipitation followed sequencing (ChIP-seq) data FLI1 histone marks from cell lines demonstrated binds a GGAA-microsatellite close CALCB, exhibits characteristics active enhancer. Reporter assays confirmed strong EWSR1-FLI1- length-dependent enhancer this GGAA-microsatellite. Mass spectrometric culture supernatants secreted cells. While short-term RNA interference-mediated had no effect on proliferation clonogenic growth cells vitro, its long-term decreased vitro vivo. Similarly, reduced clonogenic/spheroidal tumorigenicity, small-molecule inhibitors directed against RAMP1-comprising Collectively, our findings suggest direct targeting CALCB/RAMP1 axis may offer new therapeutic strategy inhibition growth.