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Cellular unjamming refers to the coordinated fluid-like motion of cells and has significant implications in various biological processes such as development, wound healing, and tumor progression. One of the contexts in which cellular unjamming becomes crucial is in tumor growth, where the unchecked proliferation of cancer cells within a confined space generates compressive stress. Since multiple cellular and molecular mechanisms are potentially operating simultaneously in this space, the predominant mechanism by which a densely packed monolayer or 3D aggregate of mechanically stressed cells can unjam is unknown. Furthermore, while cells within a multicellular aggregate are known to segregate into distinct groups based on their differential adhesive properties, the physical attributes in the tumor microenvironment that drive tumor-cell sorting and subsequent invasion remain poorly understood. First, I investigate the role of cell-cell adhesion in unjamming transitions within a mechanically stressed monolayer and establish that increased collective cell migration is correlated with cell shape and distinct from the well-known epithelial-to-mesenchymal transition (EMT). Cell-cell adhesion plays a vital role in the transition to a fluidized state, resulting in increased collective motion and accelerated wound repair while maintaining epithelial characteristics. E-cadherin knockdown inhibits coordinated migration under mechanical compression, demonstrating that the unjamming transition is not primarily driven by EMT but rather by increased cell-cell adhesions and reduced traction forces within the bulk cell sheet. The findings provide a novel perspective on tumor development and cancer invasion, suggesting that compressive stress inhibits normal epithelial cell migration but enables cancer cells to migrate rapidly as a cohesive collective. Next, I will investigate how matrix stiffness can regulate cell invasion and the sorting of two cell types with different adhesiveness into the core and periphery of 3D spheroids. ECM stiffness is known to influence cancer spread and metastasis, and I demonstrate that single-cell cancer migration is restricted by high confinement. Moreover, matrix stiffness directly impacts 3D spheroid sorting, in addition to individual and collective cell invasion. Spheroid sorting leads to high local cell density in the core, where cells strengthen cell-cell adhesions, reduce volume, and exhibit jammed behavior. In contrast, less adhesive mesenchymal cells occupy the outer edges of the spheroid in an unjammed state, enabling rearrangements within the compartment boundary. My findings show that pressure-driven burst-like collective cell motion is observed when matrix confinement is lowered for sorted spheroids, allowing rapid escape into the surrounding matrix. This has significant implications for cancer cells sorted to the tumor periphery that proceed to degrade the matrix and ultimately induce burst-driven cell invasion. Thus, my dissertation establishes compressive stress-mediated and adhesion-dependent collective cell behavior in both 2D and 3D contexts. Together, the findings shed light on the impact of compressive stress and matrix stiffness on the mechanisms of collective cell motion in cancer migration, tumor development, and distant metastasis. By improving our understanding of the mechanical inputs and forces contributing to collective migration in 2D and 3D settings, I hope that this research will lead to the development of improved therapies targeting cancer cell sorting and migration in the primary tumor.

Cell migration plays an essential role in many of physiological and pathological processes, including morphogenesis, inflammation, wound healing, and tumor metastasis. It is a complex process that involves multi-scale interactions between the cell and the extracellular matrix (ECM). Cells migrate through stromal ECM with native and cell-derived curvature at micron-meter scale are context-specific. How does the curvature of ECM mechanically change cell morphology and motility? Can the diverse migration behaviors from genetically identical cells be predictively using cell migrating data? We address these questions using an integrated computational and experimental approach: we developed three-dimensional biomechanical cell model and measured and analyzed a large number of cell migration images over time. Our findings suggest that 1. substrate curvature determines cell shape through contact and regulating protrusion dynamics; 2. effective cell migration is characterized with long cellular persistence time, low speed variation, spatial-temporally coordinated protrusion and contraction; 3. the cell shape variation space is low dimensional; and 4. migration behavior can be determined by a single image projected in the low dimensional cell shape variation space.

My research entails the design and implementation of microfluidic systems to investigate the mechanical dynamics in cancer invasion at the single -cell level. Metastasis, the process of cancer spreading, is the leading cause of cancer related deaths, but the principles and dynamics that drive this process remain largely unknown. The challenge is in part due to the large spatial and temporal scales that the metastatic process spans, as it could involve single-cells transporting to distal sites across meters and over months to years. To address this challenge, my goal is to create microsystems that aim to recapitulate the critical steps in dissemination. Cell scaled microchannels with subnucleus-scaled barriers are incorporated to elucidate the mechanical transition dynamics of invasive cancer cells. Single-cell - single-barrier interactions are induced and invasive behavior is elicited. Different regulators of invasion are explored, including molecular modulators of microtubules and actomyosin as well as mechanical factors such as dimensional, directional, and other engineered spatially asymmetric cues. Results of my work have shown that microtubule stabilization suppresses cell invasion across subnucleus-scaled barriers, physiologically ubiquitous mechanical cues such as dimensionality and directionality modulate migratory cell decision making, myosin IIa activity inhibition can alter invasion patterns, and certain mechanically asymmetric microenvironments can potentially suppress dissemination via the phenomenon iteratio ad nauseam.