Single‐Cell Morphomechanics of Prostate Cancer‐Associated Fibroblasts Identifies Distinct Features Associated with Patient Outcome

Abstract Tumor development and progression reshape the physical properties of the surrounding tumor microenvironment (TME), including its biomechanical traits. This is driven by a prominent cell type in the TME, cancer‐associated fibroblasts (CAFs), which increases tissue stiffness via extracellular matrix deposition and remodeling. Currently, it is unclear whether there are also physical changes to CAFs at the cellular level and, if so, how they relate to patient outcome. Here, it is shown that CAFs have distinct morphological and biomechanical features from normal fibroblasts. Matched, patient‐derived CAFs and non‐malignant prostate fibroblasts (NPFs) from 35 patients with primary prostate cancer are examined. Morphologically, CAFs have more aligned stress fibers and larger and more elongated nuclei, based on quantitative image analysis of confocal microscopy images. In addition, single‐cell mechanical measurements using real‐time deformability cytometry showed that CAFs are larger and stiffer than NPFs. These changes are consistent across patients and validated with atomic force microscopy. A combined morphomechanical score encompassing these features is significantly associated with patient outcome. In transcriptomic analyses, the score is correlated with microtubule dynamics and a myofibroblast phenotype. Importantly, it is also demonstrated that morphomechanical features of prostate fibroblasts are modified by approved treatments for prostate cancer, such as docetaxel, and other small molecular inhibitors, particularly those targeting FGFR. In summary, changes in cellular morphomechanical properties are a consistent feature of CAFs and are associated with patient outcome. Moreover, cellular morphomechanical properties can be therapeutically targeted, potentially providing a new strategy for manipulating the TME to control cancer progression.