Accumulating data suggest that metastatic dissemination often occurs early during tumour formation but the mechanisms of early metastatic spread have not yet been addressed. Here, we studied metastasis in a HER2-driven mouse breast cancer model and found that progesterone-induced signalling triggered migration of cancer cells from early lesions shortly after HER2 activation, but promoted proliferation in advanced primary tumour cells. The switch from migration to proliferation was regulated by elevated HER2 expression and increased tumour cell density involving miRNA-mediated progesterone receptor (PGR) down-regulation and was reversible. Cells from early, low-density lesions displayed more stemness features than cells from dense, advanced tumours, migrated more and founded more metastases. Strikingly, we found that at least 80% of metastases were derived from early disseminated cancer cells (DCC). Karyotypic and phenotypic analysis of human disseminated cancer cells and primary tumours corroborated the relevance of these findings for human metastatic dissemination.

Metastasis is the leading cause of cancer-related deaths; metastatic lesions develop from disseminated cancer cells (DCCs) that can remain dormant1. Metastasis-initiating cells are thought to originate from a subpopulation present in progressed, invasive tumours2. However, DCCs detected in patients before the manifestation of breast-cancer metastasis contain fewer genetic abnormalities than primary tumours or than DCCs from patients with metastases3–5. These findings, and those in pancreatic cancer6 and melanoma7 models, indicate that dissemination might occur during the early stages of tumour evolution3,8,9. However, the mechanisms that might allow early disseminated cancer cells (eDCCs) to complete all steps of metastasis are unknown8. Here we show that, in early lesions in mice and before any apparent primary tumour masses are detected, there is a sub-population of Her2+p-p38lop-Atf2loTwist1hiE-cadlo early cancer cells that is invasive and can spread to target organs. Intra-vital imaging and organoid studies of early lesions showed that Her2+ eDCC precursors invaded locally, intravasated and lodged in target organs. Her2+ eDCCs activated a Wnt-dependent epithelial–mesenchymal transition (EMT)-like dissemination program but without complete loss of the epithelial phenotype, which was reversed by Her2 or Wnt inhibition. Notably, although the majority of eDCCs were Twist1hiE-cadlo and dormant, they eventually initiated metastasis. Our work identifies a mechanism for early dissemination in which Her2 aberrantly activates a program similar to mammary ductal branching that generates eDCCs that are capable of forming metastasis after a dormancy phase.

Cancer cell dissemination during very early stages of breast cancer proceeds through poorly understood mechanisms. Here we show, in a mouse model of HER2+ breast cancer, that a previously described sub-population of early-evolved cancer cells requires macrophages for early dissemination. Depletion of macrophages specifically during pre-malignant stages reduces early dissemination and also results in reduced metastatic burden at end stages of cancer progression. Mechanistically, we show that, in pre-malignant lesions, CCL2 produced by cancer cells and myeloid cells attracts CD206+/Tie2+ macrophages and induces Wnt-1 upregulation that in turn downregulates E-cadherin junctions in the HER2+ early cancer cells. We also observe macrophage-containing tumor microenvironments of metastasis structures in the pre-malignant lesions that can operate as portals for intravasation. These data support a causal role for macrophages in early dissemination that affects long-term metastasis development much later in cancer progression. A pilot analysis on human specimens revealed intra-epithelial macrophages and loss of E-cadherin junctions in ductal carcinoma in situ, supporting a potential clinical relevance.

The main circulating estrogen hormone 17β-estradiol (E2) contributes to the initiation and progression of breast cancer. Estrogen receptors (ERs), as transcription factors, mediate the effects of E2. Ablation of the circulating E2 and/or prevention of ER functions constitute approaches for ER-positive breast cancer treatments. These modalities are, however, ineffective in de novo endocrine-resistant breast neoplasms that do not express ERs. The interaction of E2-ERs with specific DNA sequences, estrogen responsive elements (EREs), of genes constitutes one genomic pathway necessary for cellular alterations. We herein tested the prediction that specific regulation of ERE-driven genes by an engineered monomeric and constitutively active transcription factor, monotransregulator, provides a basis for the treatment of ER-negative breast cancer. Using adenovirus infected ER-negative MDA-MB-231 cells derived from a breast adenocarcinoma, we found that the monotransregulator, but not the ERE-binding defective counterpart, repressed cellular proliferation and motility, and induced apoptosis through expression of genes that required ERE interactions. Similarly, the monotransregulator suppressed the growth of ER-negative BT-549 cells derived from a breast-ductal carcinoma. Moreover, the ERE-binding monotransregulator repressed xenograft tumor growth in a nude mice model. Thus, specific regulation of genes bearing EREs could offer a therapeutic approach for de novo endocrine-resistant breast cancers.

No abstract

Cancer cell dissemination can occur during very early stages of breast cancer, but the mechanisms controlling this process are unclear. Here we show that a previously reported early MMTV-HER2+/P-p38lo/TWISThi/E-cadherinlo cancer cell subpopulation depends on macrophages for early dissemination. Depletion of macrophages before overt tumor detection drastically reduced early dissemination and diminished the late metastatic burden. CD206+/Tie2+ macrophages were attracted into early lesions in part by CCL2 produced by early HER2+ cancer cells and myeloid cells. Upregulation of Wnt-1 by macrophages could be stimulated by CCL2 and correlated with loss of E-cadherin in HER2+ early cancer cells. Both MMTV-PyMT and MMTV-HER2 early lesions showed macrophage-containing tumor microenvironments of metastasis (TMEM) structures, and PyMT early cancer cells also showed a reduction in early lesion E-cadherin junctions. Intraepithelial macrophages and loss of E-cadherin junctions was also found more frequently in high-grade human DCIS than in low-grade and normal breast tissue, but no association was found with HER2 status. We reveal a previously unrecognized mechanism by which macrophages play a causal role in early dissemination, impacting long-term metastasis development. Citation Format: Nina Linde, Maria Casanova-Acebes, Maria Soledad Sosa, Arthur Mortha, Adeeb Rahman, Eduardo F. Farias, Kathryn Harper, Ethan Tardio, Ivan Reyes-Torres, Joan G. Jones, John S. Condeelis, Miriam Merad, Julio A. Aguirre-Ghiso. Macrophages orchestrate early dissemination and metastasis [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2017 Oct 1-4; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2018;6(9 Suppl):Abstract nr IA16.

Metastasis is the leading cause of cancer related deaths and these lesions develop from disseminated cancer cells (DCC) that can remain dormant. Metastasis initiating cells are thought to originate from a subpopulation present in progressed invasive tumors. However, DCCs detected in patients before the manifestation of breast cancer metastasis contain fewer genetic abnormalities than primary tumors or than DCCs from patients with metastases. These findings and those in pancreatic cancer and melanoma models argued that dissemination might occur during early stages of tumor evolution. Yet, the mechanisms that might allow early-disseminated cancer cells (eDCC) to complete all steps of metastasis were unknown. Here we show that in early lesions (EL), before any overt primary tumor (PT) masses are detected, there is a sub-population of Her2+/P-p38lo/P-ATF2lo/TWISThi/E-cadherinlo early cancer cells that are invasive and disseminate to target organs. Intra-vital imaging and organoid studies of early lesions revealed that Her2+ eDCC precursors locally invaded, intravasated and lodged in target organs. Her2+ eDCCs activated a Wnt-dependent EMT-like dissemination program but without complete loss of epithelial phenotype that was reversed by Her2 or Wnt inhibition. Surprisingly, while the majority of eDCCs are TWISThi/E-cadherinlo and dormant, they eventually initiate metastasis. Our work identifies a mechanism for early dissemination whereby Her2 aberrantly activates a program similar to mammary ductal branching that spawns eDCCs capable of forming metastasis after a dormancy phase. Citation Format: Kathryn Harper, Maria Soledad Sosa, David Entenberg, Hedayatollah Hosseini, Julie Cheung, Rita Nobre, Alvaro Avivar-Valderas, Chandandaneep Nagi, Nomeda Girnius, Roger Davis, Eduardo Farias, John Condeelis, Christoph A. Klein, Julio A. Aguirre-Ghiso. Mechanism of early dissemination and metastasis in Her2+ mammary 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 3051. doi:10.1158/1538-7445.AM2017-3051

No abstract