Transcriptomic profiling of fibroblasts at different stages of lung colonization by MDA-MB-231 or MDA-MB-231-LM2 cells

Cancer-associated fibroblasts promote the development of many primary malignancies, but their function in metastatic progression is poorly understood. Here, we demonstrate that colonization of the lungs by metastatic breast cancer cells induces an inflammatory phenotype in lung fibroblasts. CXCL9 and CXCL10 are induced in an NFκB-dependent manner in metastasis-associated fibroblasts in response IL-1α and IL-1β secreted by disseminated breast cancer cells. We find that the chemokine receptor CXCR3, that binds CXCL9/10, is expressed in a small subset of breast cancer cells that exhibits greater tumor-initiating ability when co-transplanted with fibroblasts. CXCR3-expressing cancer cells maintain JNK signaling that drives IL-1A/B expression, and thus rendering this subpopulation efficient in both inducing CXCL9/10 in lung fibroblasts and responding to the chemokines. Importantly, disruption of the CXCL9/10-CXCR3 axis significantly reduces metastatic colonization in xenograft and syngeneic mouse models suggesting an essential role of this paracrine crosstalk in metastatic progression and a potential therapeutic vulnerability for the treatment of metastatic breast cancer. For profiling of fibroblasts at different metastatic stages, NSG mice (6-8 weeks of age) were injected with MDA-MB-231 (MDA) or highly metastatic MDA-MB-231-LM2 (MDA-LM2) breast cancer cells via the tail vein. At week 1 and 3 post cancer cell injection (representing micro- and macrometastatic stages, respectively), lungs from mice with similar metastatic burden were harvested. Lungs from age-matched healthy mice were used as control group. Fibroblasts were isolated from whole lungs by fluorescence-activated cell sorting (FACS) using a panel of negative selection markers and CD140a/b as positive fibroblast markers. For each time point and metastasis group, 3 biological replicates were analyzed, and each biological replicate encompasses 1-3 mice.