Abstract 178: MYO10 aberrant methylation and overexpression in leader cells regulates lung cancer collective cell invasion and fibronectin patterning

Abstract

Abstract Myosin-X (MYO10) is a noncanonical myosin that drives filopodia formation and extension, and is necessary for invasion and metastasis of breast cancer, prostate cancer and melanoma. It is not yet known by what mechanism MYO10 increases cancer cell invasion and metastasis. Two independently published MYO10 knockout mouse models showed severe developmental defects dependent upon collective migration of several cell types, suggesting that MYO10 may also regulate collective invasion of cancer cells. In order to study collective cancer cell invasion, our lab previously developed a technique to isolate and culture purified highly-invasive leader cells and highly-proliferative follower cells from collectively invading lung cancer cell lines. Using this method, we demonstrated distinct gene expression patterns, DNA methylation patterns, and phenotypic properties of leader versus follower cells. MYO10 is highly overexpressed in leader cells but is not expressed in follower cells, and MYO10 overexpression in leaders correlates with highly significant CpG island promoter DNA hypomethylation and gene body DNA hypermethylation. We hypothesize that MYO10 regulates cancer cell collective invasion by driving the formation of long filopodia necessary for leading-edge fibronectin patterning and subsequent leader cell invasion. MYO10 localizes at leader cell filopodia tips but not in follower cell filopodia within both 2D culture and 3D invading spheroids of four non-small cell lung cancer cell lines. MYO10 knockdown in purified leader cells and in these four lung cancer cell lines decreases the number and length of filopodia, 2D cell motility and 3D spheroid invasion; in contrast, MYO10 overexpression in follower cells increases filopodia length, cell motility and spheroid invasion. In addition, proteomic analysis shows that leader cells produce and secrete fibronectin (FN1), unlike follower cells. Immunofluorescence within invading spheroids shows the formation of long FN1 fibrils, i.e. elongated parallel bundles of FN1 that extend far past the leader cell body in multiple cell lines. FN1 fibrils preferentially localized with MYO10+ filopodia. Knockdown of MYO10 disrupts the formation of these fibronectin fibrils. We previously reported that knockdown of fibronectin completely abrogates lung cancer spheroid collective invasion. Therefore, our data suggest that MYO10 regulates cancer cell collective invasion by driving the formation of long filopodia that then regulate fibronectin architecture and subsequent invasion by leader cells. Note: This abstract was not presented at the meeting. Citation Format: Emily R. Summerbell, Jessica Konen, Jeanne Kowalski, Paula Vertino, Adam Marcus. MYO10 aberrant methylation and overexpression in leader cells regulates lung cancer collective cell invasion and fibronectin patterning [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 178.