Tumor-associated macrophages drive stromal cell-dependent collagen crosslinking and stiffening to promote breast cancer aggression

Stromal stiffening accompanies malignancy, compromises treatment, and promotes tumor aggression. Clarifying the molecular nature and the factors that regulate stromal stiffening in tumors should identify biomarkers to stratify patients for therapy and interventions to improve outcome. We profiled lysyl hydroxylase- and lysyl oxidase-mediated collagen crosslinks and quantified the greatest abundance of total and complex collagen crosslinks in aggressive human breast cancer subtypes with the stiffest stroma. These tissues harbor the highest number of tumor-associated macrophages (TAM), whose therapeutic ablation in experimental models reduced metastasis, and decreased collagen crosslinks and stromal stiffening. Epithelial-targeted expression of the crosslinking enzyme, lysyl oxidase, had no impact on collagen crosslinking in PyMT mammary tumors, whereas stromal cell targeting did. Stromal cells in microdissected human tumors expressed the highest level of collagen crosslinking enzymes. Immunohistochemical analysis of a cohort of breast cancer patient biopsies revealed that stromal expression of lysyl hydroxylase two, an enzyme that induces hydroxylysine aldehyde-derived collagen crosslinks and stromal stiffening, correlated significantly with disease specific mortality. The findings link tissue inflammation, stromal cell-mediated collagen crosslinking and stiffening to tumor aggression and identify lysyl hydroxylase two as a stromal biomarker.