Tumor growth generates mechanical compressive stress within the tumor mass, which drives tumor cells to undergo mesenchymal-amoeboid transition (MAT). MAT tumor cells adopt fast amoeboid migration to traverse confined microenvironments such as extracellular matrix gaps and blood vessel walls, mediating distant tumor metastasis. However, the cell fate of these MAT tumor cells under persistent strong confinement remains unclear. The canonical view holds that MLKL, the executioner protein of necroptosis, requires phosphorylation by the upstream kinase RIPK3 for activation. Nevertheless, recent studies have detected necroptosis in human tumors lacking RIPK3, and the molecular mechanism underlying this phenomenon remains elusive.
In a study published in Developmental Cell, a research team led by Prof. SUN Liming from the Center for Excellence in Molecular Cell Science (Shanghai Institute of Biochemistry and Cell Biology) of the Chinese Academy of Sciences, in collaboration with Prof. LIU Yan-Jun from the Institutes of Biomedical Sciences, Fudan University, reveals that upon persistent physical confinement, tumor cells that undergo mesenchymal-amoeboid transition (MAT) activate an RIPK3-independent, MLKL-mediated programmed necroptosis pathway. Activation of this pathway not only induces cell death but also suppresses amoeboid migration of tumor cells, serving as an endogenous tumor-suppressive checkpoint. Under the selective pressure of tumor evolution, escape clones with low MLKL expression are selected, which drive tumor metastasis with enhanced migratory and invasive capacities.
In this study, the researchers established an in vitro L-MAT (Long-term Mesenchymal-amoeboid Transition) experimental system to mimic the compressive microenvironment in tumors. They found that sustained mechanical compression directly upregulates MLKL transcription; once MLKL expression exceeds the tolerance threshold of tumor cells, RIPK3-independent necroptosis is triggered. Further mechanistic dissection demonstrates that the mechanosensitive ion channel PIEZO1 senses compressive signals and directly drives MLKL transcription via the downstream transcription factor STAT1. The accumulated MLKL protein translocates to the cell membrane and subsequently initiates necroptosis.
Notably, MLKL activation suppresses amoeboid migration of MAT tumor cells prior to plasma membrane rupture and necrotic cell death. Activated MLKL disrupts retrograde cortical actin flow and downregulates the expression of migration-related genes, ultimately halting tumor cell motility. Upon MLKL knockout, tumor cells evade this cell-death checkpoint and retain fast amoeboid migratory properties. A series of in vitro and in vivo models including Transwell migration assays, microfluidic chips and mouse lung metastasis models collectively demonstrate that MLKL activation markedly impairs the invasion and metastasis of tumor cells within confined microenvironments.
The researchers further analyzed spatial transcriptomic data from clinical tumor samples. In human breast cancer specimens, MLKL expression is high and the PIEZO1-STAT1 pathway is activated in less malignant in situ carcinoma regions, accompanied by abundant CD8⁺ T cell infiltration. In contrast, MLKL expression is markedly downregulated in highly invasive invasive carcinoma regions, where the PIEZO1-STAT1 axis becomes uncoupled and anti-tumor immune infiltration is diminished. In clinical samples of colorectal cancer liver metastasis, MLKL expression is significantly lower in liver metastatic lesions compared with primary tumors. These findings suggest that under tumor evolutionary pressure, tumor clones with low MLKL expression and necroptosis resistance are enriched. These clones escape both cell death and immune surveillance, gain enhanced migratory capacity, and ultimately promote malignant progression and distant metastasis.
In summary, this study defines the fate of MAT tumor cells under persistent confinement: these cells undergo RIPK3-independent, MLKL-mediated necroptosis. The PIEZO1-STAT1-MLKL pathway functions as a mechanosensory checkpoint to eliminate highly invasive amoeboid tumor cells; tumor evolution selects for MLKL-low escape clones to facilitate tumor invasion and metastasis. This work expands our understanding of the activation modes of necroptosis and provides important theoretical foundations for discovering tumor prognostic biomarkers and developing novel anti-metastatic therapeutic targets.
This finding connects directly to what happens in our bodies every day. Cells constantly experience mechanical forces as part of normal tissue homeostasis. However, during solid tumor development, cancer cells proliferate and migrate within confined spaces and are subjected to progressively increasing mechanical pressure. This study suggests that cells can interpret this abnormal physical environment as a danger signal.
Reference: https://doi.org/10.1016/j.devcel.2026.09.003
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