SRA

Dormancy of disseminated tumor cells in secondary organs leads to late breast cancer-related deaths after relapse. The bone marrow is one of the primary breast cancer metastatic sites and is associated with poor prognosis. Here, we investigated the role of major factors of the bone marrow, BMP4 and TGF?2, in regulating cancer cell dormancy. Unexpectedly, we observed that TGF?2 and BMP4 have a synergistic effect that induces dormancy in both normal and transformed mammary stem cells. Several assays (3D matrigel, FUCCI Cell Cycle Indicator) showed that co-exposure to TGF?2 and BMP4 had a stronger anti-proliferative effect than each ligand alone. In addition, transformed cells fully retained this synergistic effect while they became less sensitive to individual cytokines. Surprisingly, single-cell RNAseq analysis revealed the heterogeneity of the G0 compartment at the transcriptomic level. We identified a unique deep dormant cluster under TGF?2 and BMP4 co-exposure characterized by a blended signature from treatments by TGF?2 or BMP4 alone. These findings reveal that disseminated breast cancer cells in the bone marrow are placed in a deep dormant stage by the synergistic effect of TGF?2 and BMP4 that neither factor can achieve alone. Lastly, our data suggest that the local BMP4 levels decrease with tissue aging and can therefore contribute to dormant cell awakening. By providing a better understanding of BMP4/TGF?2 signaling, our results open opportunities to prevent cancer relapse. Overall design: We performed RNAseq to assess the differential programs inducing dormancy by TGFB2 and/or BMP4 treatment PYMT cells were treated with 5ng/mL of TGF?2, BMP4, or both for 3 days.