Tumor matrix stiffness drives malignant progression in murine breast cancer: enhanced stemness, tumorigenesis and metastasis

Abstract

Matrix stiffness in the tumor microenvironment influences cancer cell behavior, including tumor growth and metastasis. We tested whether prior exposure to different stiffness levels creates a lasting mechanical memory that affects subsequent tumorigenesis and metastatic potential. We cultured 4T1 murine breast cancer cells on alginate–chitosan scaffolds mimicking normal (0.51 ± 0.13 kPa), fibrotic (3.40 ± 0.53 kPa), or tumoral (11.40 ± 0.96 kPa) breast tissue stiffness. We assessed cell morphology, proliferation, and expression of stemness markers (Sox2, Nanog, Oct4). After detachment, cells were injected into mouse mammary fat pads, and tumor latency, volume, and weight were monitored for 30 days. Lung and liver tissues were examined histologically for metastatic lesions and Plac1 mRNA levels. Cells preconditioned on the stiff, tumor-mimicking scaffolds showed increased proliferation and stemness-marker expression in vitro and developed tumors with shorter latency, larger volume and weight, and more lung and liver metastases in vivo. These results indicate that a stiff matrix can imprint a persistent, pro-metastatic phenotype, consistent with a model of mechanical memory, enhancing tumorigenicity and metastatic spread. Targeting the mechanisms underlying this memory may offer new strategies to prevent breast cancer progression.