Updated project metadata. MYCN amplification (MNA) is a defining feature of high-risk neuroblastoma (NB) that predicts poor prognosis. However, whether genes within or in close proximity to the MYCN amplicon also contribute to aggressiveness in MNA+ NB remains poorly understood. Here we identify that GREB1, a transcription factor encoding gene neighboring the MYCN locus, is frequently co-expressed with MYCN, and promotes cell survival in MNA+ NB. GREB1 controls gene expression independently of MYCN in MNA+ NB, among which we uncover Myosin 1B (MYO1B) as being highly expressed in MNA+ NB. MYO1B promotes aggressive features, including invasive capacity in vitro, as well as extravasation and distant metastasis in vivo. Global secretome and proteome profiling further delineate MYO1B as a major regulator of secretome reprogramming in MNA+ NB cells. Moreover, we identify the cytokine MIF as an important pro-invasive and pro-metastatic mediator of MYO1B activity. Together, we have identified a putative GREB1-MYO1B-MIF axis as an unconventional mechanism that promotes the aggressiveness of MNA+ NB, and independently of MYCN. Furthermore, we find that MYO1B is upregulated in association with other oncoproteins during cellular transformation, and is dramatically increased in multiple human cancer types, suggesting a crucial role of MYO1B in cancers in addition to MNA+ NB.