Abstract P4-03-18: A hierarchy of cancer associated fibroblasts in situ and in circulation promote breast cancer metastasis
Authors: Sharma U , Miller P , Speransky S , Medina-Saenz K , Ferrer P , Lippman M , El-Ashry D 
Published: March 2017

Background: Metastasis is the primary cause of breast cancer mortality. Interactions between cancer cells and non-cancer cells of the tumor microenvironment (TME) are pivotal in governing tumor initiation, progression and metastasis, and cancer associated fibroblasts (CAFs) are critical orchestrators of these interactions. We recently identified circulating CAFs (cCAFs) as a novel circulating biomarker associated with metastatic breast cancer. We established CAF cell lines from dissociated luminal A, ER- Her-2 amplified, and triple-negative/basal-like (TN) breast tumors. We demonstrated that “aggressive” CAFs differentially secrete miRNAs that contribute to ER-negativity, activated growth factor signaling, and induction of EMT in breast cancers compared to “indolent” CAFs. We hypothesized that a hierarchy exists within CAFs regarding their ability to facilitate tumor progression and metastasis. Here we demonstrate that CAFs derived from aggressive TN breast tumors differ from those derived from more indolent Luminal A breast tumors in secretion of cytokines and chemokines that can confer differential effects on the behavior of breast cancer cells. We also demonstrate that “aggressive” CAFs more potently facilitate tumor progression and metastasis than “indolent” CAFs. We additionally evaluated if “aggressive” and “indolent” CAFs differ in their ability to mobilize CTCs and circulating CAFs into circulation.

Methods: Conditioned media (CM) from “aggressive” and “indolent” CAFs was analyzed for chemokine/cytokine expression. Luminal A breast cancer cells (MCF-7) or primary tumor cells from an aggressive TN tumor (DT28) were injected into the mammary fat pad of 6-8 week old female NSG mice, either alone or in combination with CAF19-I or CAF23-A. Tumor progression was monitored and mice were examined for metastasis at necropsy. Tissues were harvested for histology and blood was collected by cardiac puncture. Plasma was analyzed for cytokine/chemokine expression and blood was processed for enumeration of circulating tumor cells (CTCs) and cCAFs.

Results: “Aggressive” CAF CM had significantly higher levels of a number of factors, including IL-8, SDF-1, and CXCL1, compared to “indolent” CAF CM. MCF-7 cells co-injected with “aggressive” CAFs formed tumors much faster than those co-injected with the “indolent” CAFs or without CAFs. While DT28 cells readily form tumors and metastasize in the NSG model, fewer DT28 cells do not form metastases in the timeframe that this same lower number of DT28 cells co-injected with “aggressive” CAFs demonstrated robust tumor growth and developed metastases in liver and pancreas. DT28 cells co-injected with “indolent” CAFs did not exhibit metastases.

Conclusion: The data presented here further demonstrate that there is a hierarchy within CAFs regarding their ability to facilitate tumor growth and metastasis, and that this may largely be mediated by secreted soluble factors. “Aggressive” CAFs may retain their programmed role in circulation and accelerate metastasis more than “indolent” CAFs. We suggest that targeting CAFs in situ and in circulation and disrupting their interactions with breast cancer cells could provide novel strategies to combat breast cancer and breast cancer metastasis.

Citation Format: Sharma U, Miller P, Speransky S, Medina-Saenz K, Ferrer P, Lippman M, El-Ashry D. A hierarchy of cancer associated fibroblasts in situ and in circulation promote breast cancer metastasis [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P4-03-18.