Phenotypic plasticity describes the ability of cancer cells to undergo dynamic, nongenetic cell state changes that amplify cancer heterogeneity to promote metastasis and therapy evasion. Thus, cancer cells occupy a continuous spectrum of phenotypic states connected by trajectories defining dynamic transitions upon a cancer cell state landscape. With technologies proliferating to systematically record molecular mechanisms at single-cell resolution, we illuminate manifold learning techniques as emerging computational tools to effectively model cell state dynamics in a way that mimics our understanding of the cell state landscape. We anticipate that “state-gating” therapies targeting phenotypic plasticity will limit cancer heterogeneity, metastasis, and therapy resistance. Significance: Nongenetic mechanisms underlying phenotypic plasticity have emerged as significant drivers of tumor heterogeneity, metastasis, and therapy resistance. Herein, we discuss new experimental and computational techniques to define phenotypic plasticity as a scaffold to guide accelerated progress in uncovering new vulnerabilities for therapeutic exploitation.

Matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) of metabolites can reveal how metabolism is altered throughout heterogeneous tissues. Here negative ion mode MALDI-MSI has been coupled with laser post-ionisation (MALDI-2) and applied to the MSI of low molecular weight (LMW) metabolites (<m/z 600) to investigate the benefits MALDI-2 offers for spatial metabolomics in terms of metabolite coverage and sensitivity. When applied to mouse kidney tissue MALDI-2 provided almost double the number of on-tissue specific mass features compared to conventional MALDI. MALDI-2 also resulted in not only the increased detection sensitivity for multiple metabolite species but also permitted the imaging of LMW metabolites (e.g. uridine) that were not detected using conventional MALDI-MSI. When compared against ∼140 publically available kidney datasets submitted through the METASPACE analysis platform using the same N-(1-naphthyl) ethylenediamine dihydrochloride (NEDC) matrix, MALDI-2 provided 34 unique metabolite m/z features that were not consistently annotated previously. To further evaluate the usefulness of this MALDI-2 approach to metabolite imaging, MALDI-2 was applied to the imaging of mouse liver tissue containing a metastasised breast cancer at a pixel size of 20 μm. Using a co-localisation analysis, MALDI-2 detected six tumour-specific metabolites that were not detected using conventional MALDI, as well as providing an up to 20-fold increase in signal intensities for many others (e.g., glutamate). This work provides one of the first reports of MALDI-2 applied to metabolite imaging and demonstrates the dramatic improvements in sensitivity and metabolite coverage it provides.

Therapies that prevent metastatic dissemination and tumor growth in secondary organs are severely lacking. A better understanding of the mechanisms that drive metastasis will lead to improved therapies that increase patient survival. Within a tumor, cancer cells are equipped with different phenotypic and functional capacities that can impact their ability to complete the metastatic cascade. That phenotypic heterogeneity can be derived from a combination of factors, in which the genetic make-up, interaction with the environment, and ability of cells to adapt to evolving microenvironments and mechanical forces play a major role. In this review, we discuss the specific properties of those cancer cell subgroups and the mechanisms that confer or restrict their capacity to metastasize.

Metastasis is responsible for most cancer-related deaths. However, only a fraction of circulating cancer cells succeed in forming secondary tumours, indicating that adaptive mechanisms during circulation play a part in dissemination. Here, we report that constriction during microcapillary transit triggers reprogramming of melanoma cells to a tumorigenic cancer stem cell-like state. Using a microfluidic device mimicking physiological flow rates and gradual capillary narrowing, we show that compression through narrow channels causes cell and nuclear deformation, rapid chromatin remodelling and increased calcium signalling via mechanosensor PIEZO1. Within minutes, cells upregulate transcripts associated with metabolic reprogramming and metastatic processes. Over time, this results in the stable adoption of a cancer stem cell-like state. Squeezed cells express elevated melanoma stem cell markers, exhibit increased trans-endothelium invasion and display enhanced tumorigenicity in vitro and in vivo. Pharmacological inhibition of PIEZO1 blocks this transition, while activation with Yoda1 induces the stem cell-like state irrespective of constriction. Deletion of PIEZO1 completely abolishes the constriction-induced phenotype. Together, these findings demonstrate that compressive forces during circulation reprogram circulating cancer cells into tumorigenic, stem cell-like states, primed for extravasation and metastatic colonization.