Transcriptomic profiling of CXCL9/10-overexpressing (OE) MDA-MB-231 breast cancer cells upon treatment with CXCR3i or DMSO vehicle

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 CXCR3 inhibitor (CXCR3i)-treated or vehicle-treated breast cancer cells, 500,000 MDA-MB-231 (MDA) cells overexpressing CXCL9 and CXCL10 were seeded into ultra-low attachment cell culture flasks in 20 ml Onco2 medium, consisting of HuMEC-medium (Invitrogen) supplemented 50 U/ml penicillin (Sigma-Aldrich), 50 μg/ml streptomycin (Sigma-Aldrich), 10 ng/ml basic fibroblast growth factor (bFGF, Invitrogen), 20 ng/ml EGF (Sigma-Aldrich), 5 μg/ml human insulin (Sigma-Aldrich) and 2 % vol/vol B27 (Life Technologies) and incubated at 37 °C for 7 days. Cells were stimulated with 1 µM AMG-487 or 0.001% DMSO vehicle daily for 7 days. RNA from three biological replicates was analyzed using Affymetrix Gene Chip human U133 Plus 2.0 Arrays.