Multiple juxtacrine and paracrine interactions occur between cancer cells and non-cancer cells of the tumor microenvironment (TME) that direct tumor progression. Cancer Associated Fibroblasts (CAFs) are an integral component of the TME, and the majority of breast tumor stroma is comprised of CAFs. Heterotypic interactions between cancer cells and non-cancer cells of the TME occur via soluble agents, including cytokines, hormones, growth factors, and secreted microRNAs. We previously identified a microRNA signature indicative of hyperactive MAPK signaling (hMAPK-miRNA signature) that significantly associated with reduced recurrence-free and overall survival. Here we report that the hMAPK-miRNA signature associates with a high metric of stromal cell infiltrate, and we investigate the role of microRNAs, particularly hMAPK-microRNAs, secreted by CAFs on estrogen receptor (ER) expression in breast cancer cells. ER-positive MCF-7/ltE2- cells were treated with conditioned media (CM) from CAFs derived from breast cancers of different PAM50 subtypes (CAFBAS, CAFHER2, and CAFLA). CAF CM isolated specifically from ER-negative primary breast tumors led to ER repression in vitro. Nanoparticle tracking analysis and transmission electron microscopy confirmed the presence of CAF-secreted exosomes in CM and the uptake of these exosomes by the ER+ MCF-7/ltE2- cells. Differentially expressed microRNAs in CAF CM as well as in MCF-7/ltE2- cells treated with this CM were identified. Knockdown of miR-221/222 in CAFBAS resulted in knockdown of miR221/222 levels in the conditioned media and the CM from CAFBAS; miR221/222 knockdown rescued ER repression in ER-positive cell lines treated with CAFBAS-CM. Collectively, our results demonstrate that CAF-secreted microRNAs are directly involved in ER-repression, and may contribute to the MAPK-induced ER repression in breast cancer cells.
Bibliographical noteFunding Information:
The authors would like to thank Dr. Marc E. Lippman and members of Dr. El-Ashry and Dr. Lippman laboratory group, Drs. Bal Lokeshwar, Kerry Burnstein, Ram Datar and Sharon Elliot for thoughtful discussions. Sanket H. Shah would like to thank partial support and assistance from the Sheila and David Fuente Graduate Program in Cancer Biology, University of Miami. Marta Garcia-Contreras would like to thank partial support and assistance from School of Medicine and Dentistry, Catholic University of Valencia, Valencia, Spain and Ri.Med Foundation, Palermo, Italy. We thank Sylvester Comprehensive Cancer Center development funds for partial support to DEA. We would like to thank Sion Williams at Oncogenomics Core Facility of University of Miami for their technical support with the Nanostring analysis. We would like to acknowledge Mario Soriano-Navarro for his technical support at the Electron Microscopy Service of Centro de Investigación Príncipe Felipe, Valencia, Spain. Also we thank the Microvesicles and Exosomes core at Diabetes Research Institute, University of Miami for the nano-particle tracking analysis technical support. And Maria Boulina for technical support at the Analytical Imaging Core Facility core of University of Miami.
© 2015 Taylor & Francis Group, LLC.
- Estrogen receptor