Dynamic plasticity within the EMT spectrum, rather than static mesenchymal traits, drives tumor heterogeneity and metastatic progression of breast cancers

Abstract

The Epithelial-to-Mesenchymal Transition (EMT) is a developmental cellular program frequently coopted by cancer cells and is a key contributor to both heterogeneity in solid tumors and later stage chemo-resistance and metastasis. Rather than being a switch from an epithelial to a mesenchymal state, increasing evidence points to the existence of intermediate EMT states, wherein cells co-express both epithelial and mesenchymal traits. Multiple stable intermediate EMT states possessing unique characteristics exist across the EMT spectrum, contributing to the complex heterogeneity of tumors to promote metastasis16. While much work has been carried out identifying and characterizing EMT-inducing transcription factors, the transcriptional and epigenetic networks responsible for the stability and maintenance of the midpoints along the EMT spectrum are poorly defined. In addition, there are currently no approaches to identifying and quantifying intermediate EMT subpopulations within patient tumors to evaluate their prognostic significance. Using the SUM149PT breast cancer cell line, we systematically interrogate how each EMT state independently contributes to heterogeneity and influences metastatic progression, uncovering the role of RUNX2 in stabilizing certain intermediate states. Using SUM149PT-derived tumors as a training set, we develop an entropy-based model to quantify phenotypic heterogeneity and EMT status. Remarkably, the majority of cell states captured in the SUM149PT model are represented in a selection of patient tumors, laying the foundation for quantification of epithelial-mesenchymal heterogeneity and understanding the role of the intermediate EMT state in tumor progression.

Distinct Cytosine Modification Profiles Define Epithelial-to-Mesenchymal Cell-State Transitions

“…Previous work establishing this model system has demonstrated that the intermediate clones represent a population of tumor cells with high tumor-initiating potential(8). We show high 5hmC CpGs association with motifs of key EMT transcription factors which may indicate recruitment of various transcription factors by 5hmC may be a potential mechanism regulating the intermediate clones' high tumor-initial potential.…”

“…To explore potential functional differences in gene regulation between intermediate and distal clones, we identi ed regions of accessible chromatin using ATAC-seq and conducted transcription factor motif enrichment analysis of accessible chromatin peaks containing at least one high 5hmC CpG each for intermediate clones (8,975 CpGs in 2,616 chromatin accessible peaks) and distal clones (21,022 CpGs in 3,572 chromatin accessible peaks). Motif enrichment analysis found 409 transcription factors (TF) signi cantly associated with high 5hmC chromatin accessible peaks in intermediate clones compared to 29 TFs in distal clones under the FDR < 0.05 threshold (Fig.…”

“…Cells that reside in an intermediate state display more plasticity than the cells on either ends of the EMT spectrum (3,(6)(7)(8). In addition to increased plasticity, intermediate cells have been shown to harbor stem cell characteristics such as self-renewal and increased expression of pluripotent genes (9)(10)(11).…”

Genomic events shaping epithelial-to-mesenchymal trajectories in cancer

“…Moreover, stable late hEMT states have been linked with maximal metastatic potential and worse overall survival 55 . Undoubtedly, the hEMT state can be further subdivided into sub-states, as shown by Goetz et al 4 , Brown et al 56 and also by us in the tissue-specific analyses. The true number of EMT intermediate states is just beginning to be explored.…”

Genomic events shaping epithelial-to-mesenchymal trajectories in cancer

“…Moreover, stable late hEMT states have been linked with maximal metastatic potential and worse overall survival 55 , explaining the variable metastatic potential we observed in cell lines in the context of hEMT. Undoubtedly, the hEMT state can be further subdivided into sub-states, as shown by Goetz et al 4 , Brown et al 56 and also by us in the tissue-specific analyses. The true number of EMT intermediate states is just beginning to be explored.…”