Abstract 5895: Circulating CAF and cCAF circulating tumor cell co clusters are associated with metastatic breast cancer
Authors: Sharma Utsav , Miller Philip , Medina-Saenz Kelsie , Ferrer Pedro , Speransky Svetlana , Yeasky Toni , El-Ashry Dorraya 
Published: July 2017

Abstract
Background: Breast cancer metastasis is the cause of breast cancer-related mortality. The tumor microenvironment (TME) plays a critical role in governing tumor initiation, progression and metastasis. In breast cancer, cancer associated fibroblasts (CAFs) are master regulators of the TME. Cancer metastasis occurs in part by transport of circulating tumor cells (CTCs) in the circulation. CTCs in clusters, rather than individual CTCs, have a greater capacity to establish metastases. We recently demonstrated that CAFs also circulate (cCAFs), both in clusters with CTCs and individually, in the blood of the majority of breast cancer patients with metastases, some patients without overt metastases, and in no patients with no evidence of disease. Our lab has also established primary breast cancer and primary CAF cell lines from dissociated breast tumors of different molecular subtypes; these are ideal models to dissect tumor-stromal interactions, both in vitro and in vivo. We hypothesize that cCAFs originate from the primary tumor, and further, that cCAFs cluster with CTCs to facilitate metastatic seeding.
Methods: We used separately labeled CAFs and primary breast cancer cells (DT28) co-injected into NSG mice and followed by a novel 2-color IVIS. We also modeled the ability of CAFs and breast cancer cells to form clusters in culture and used our microfluidic filter technology to evaluate the composition of clusters resulting from admixed CAFs and BC cells of differing metastatic capacity – metastatic DT28 and non-metastatic MCF-7.
Results: Through sequential dual-color IVIS we demonstrate that we can monitor the presence of CAFs and breast cancer cells at orthotopic tumor injection sites and to sites of distant metastasis, indicating that cCAFs originate from the primary tumor. Evaluation of cCAFs and CTCs in tumor bearing mice indicates that breast cancer cells with high metastatic potential mobilize greater numbers of cCAFs. CAFs formed robust clusters with metastatic DT28 cells, while non-metastatic MCF-7 cells clustered with each other but not with CAFs suggesting that the ability of breast cancer cells to form clusters with CAFs, both in vitro and in vivo, is reflective of the metastatic capability of the breast cancer cell line.
Conclusions: The intrinsic metastatic capabilities of breast cancer cells are augmented by contact and clustering with cCAFs. In turn, the ability of CAFs to mobilize into circulation is in part conferred by properties of aggressive breast cancer cells. These data corroborate observations from our pilot clinical study that indicated the presence of cCAFs is overwhelmingly associated with the presence of metastatic breast cancer.
Citation Format: Utsav Sharma, Philip Miller, Kelsie Medina-Saenz, Pedro Ferrer, Svetlana Speransky, Toni Yeasky, Dorraya El-Ashry. Circulating CAF and cCAF circulating tumor cell co clusters are associated with metastatic breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5895. doi:10.1158/1538-7445.AM2017-5895