Novel biomarkers in mutation-specific gene profiles highlight the involvement of COL9A3 in cancer cell plasticity in uveal melanoma

UM is a deadly ocular malignancy with well-described genetic alterations that predict disease outcome. However, our current understanding of the biological underpinnings of high-risk uveal melanoma progression remains relatively limited. On the basis of RNA expression profiles, we identified 12 novel biomarkers associated with high-risk UM, with protein expression validation of the 2 top markers RBFOX2 and COL9A3. Moreover, we investigated the functional contribution of COL9A3 via its overexpression in low-risk and high-risk UM cell lines. Our data suggest that COL9A3 enhances cell motility and cell proliferation and alters morphology specifically in high-risk UM cells. Furthermore, zebrafish xenograft studies revealed improved cell dissemination in a low-risk UM cell line upon overexpression of COL9A3, whereas a high-risk UM cell line remained under wild-type conditions. Interestingly, RNA sequencing revealed a high-stress profile in high-risk UM cell lines overexpressing COL9A3 and suggested the upregulation of plasticity markers, such as CD44, Nestin, EZH2, ABCB5, PAX3 and CD166, as a coping mechanism. These markers are known as differentiation markers during the development of melanocyte biogenesis and are implicated in plasticity in other high-risk cancers. The relationship between COL9A3 and plasticity was validated by multiplex immunohistochemistry of FFPE tissue, which demonstrated colocalization of COL9A3, CD44 and Nestin. BAP1mut-UM samples presented higher levels of COL9A3, CD44 and Nestin expression than EIF1AXmut- or SF3B1mut UM samples did. A positive correlation between COL9A3 and Nestin expression in triple-positive cells, regardless of the UM subtype, was observed. Together, our data suggest that COL9A3 is a novel high-risk biomarker that may be involved in UM cell plasticity and is most prevalent in BAP1mut UM.