The cytoskeleton is a participant in key cell events in oncogenesis and the reaction of cancer cells to therapeutic factors. The state and structure of the actin cytoskeleton contributes significantly to the mechanical behavior of cancer cells, forming the main features of their mechanical phenotype.In the work, using atomic-force microscopy and fluorescent microscopy, we studied the changes in the structure of the actin cytoskeleton and parameters of the mechanical properties of breast cancer cells of different molecular biological subtypes (hormone-sensitive, line ZR-75, and triple-negative, BT-20 line, subtypes) when cells interact with the CD109 antigen, a TGF-β signaling pathway inhibitor. The use of antibodies for immobilizing the CD109 antigen has been shown to cause significant changes in the spatial organization of the actin cytoskeleton, stiffness and adhesive properties of the cell surface of both cell lines. Because of differences in the structure of the actin cytoskeleton, changes in the mechanical properties of the cells of different molecular biological breast cancer subtypes and the implementation of the TGF-β signaling pathway in these cells when binding to antibodies against the CD109 antigen occur in different ways. The obtained data open new perspectives for the development and evaluation of the effectiveness of anticancer drugs.