Computational modeling reveals a vital role for proximity-driven additive and synergistic cell-cell interactions in increasing cancer invasiveness.

Solid-tumor cell invasion typically occurs by collective migration of attached cell-cohorts, yet we show here that indirect cell-interactions through the substrate can also drive invasiveness. We have previously shown that well-spaced, invasive cancer cells push-into and indent gels to depths of 10 µm, while closely adjacent, non-contacting cancer cells may reach up to 18 µm, potentially relying on cell-cell interactions through the gel-substrate. To test that, we developed finite element models of indenting cells, using experimental gel mechanics, cell mechanostructure, and force magnitudes. We show that under 50-350 nN of combined traction and normal forces, a stiff nucleus-region is essential in facilitating 5-10 µm single-cell indentations, while uniformly soft cells attain 1.6-fold smaller indentations. We observe that indentation depths of cells in close proximity (0.5-50 µm distance) increase relative to well-spaced cells, due to additive, continuum mechanics-driven contributions. Specifically, 2-3 cells applying 220 nN normal forces gained up to 3% in depth, which interestingly increased to 7.8% when two cells, 10 µm apart, applied unequal force-magnitudes (i.e., 220 and 350 nN). Such additive, energy-free contributions can reduce cell mechanical energy -output required for invasiveness, yet the experimentally observed 10-18 µm depths likely necessitate synergistic, mechanobiological changes, which may be mechanically triggered. We note that nucleus stiffening or cytoplasm softening by 25-50% increased indentation depths by only 1-7%, while depths increase nearly linearly with force-magnitude even to two-fold levels. Hence, cell-proximity triggered, synergistic and additive cell-interactions through the substrate can drive collective cancer-cell invasiveness, even without direct cell-cell interactions. STATEMENT OF SIGNIFICANCE: Metastatic cancer invasion typically occurs collectively in attached cell-cohorts. We have previously shown increased invasiveness in closely adjacent cancer cells that are able to push-into and indent soft-gels more deeply than single, well-spaced cells. Using finite element models, we reveal mechanisms of cell-proximity driven invasiveness, demonstrating an important role for the stiff nucleus. Cell-proximity can additively induce small increase in indentation depth via continuum mechanics contributions, especially when adjacent cells apply unequal forces, and without requiring increased cell-mechanical-energy-output. Concurrently, proximity-triggered synergistic interactions that produce changes in cell mechanics or capacity for increased force-levels can facilitate deep invasive-indentations. Thus, we reveal concurrent additive and synergistic mechanisms to drive collective cancer-cell invasiveness even without direct cell-cell interactions.