Cell lineage tracing links ERα loss in Erbb2-positive breast cancers to the arising of a highly aggressive breast cancer subtype

Abstract

HER2-positive (HER2+) breast cancers (BrCs) contain approximately equal numbers of ERα+HER2+ and ERα−HER2+ cases. An enduring obstacle is the unclear cell lineage-related characteristics of these BrCs. Although ERα+HER2+ BrCs could lose ERα to become ERα−HER2+ BrCs, direct evidence is missing. To investigate ERα dependencies and their implications during BrC growth and metastasis, we generated ERαCreRFP-T mice that produce an RFP-marked ERα+ mammary gland epithelial cell (MGEC) lineage. RCAS virus-mediated expression of Erbb2, a rodent Her2 homolog, first produced comparable numbers of ERα+RFP+Erbb2+ and ERα−RFP−Erbb2+ MGECs. Early hyperplasia developed mostly from ERα+RFP+Erbb2+ cells and ERα−RFP−Erbb2+ cells in these lesions were rare. The subsequently developed ductal carcinomas in situ had 64% slow-proliferating ERα+RFP+Erbb2+ cells, 15% fast-proliferating ERα−RFP+Erbb2+ cells derived from ERα+RFP+Erbb2+ cells, and 20% fast-proliferating ERα−RFP−Erbb2+ cells. The advanced tumors had mostly ERα−RFP+Erbb2+ and ERα−RFP−Erbb2+ cells and only a very small population of ERα+RFP+Erbb2+ cells. In ERα−RFP+Erbb2+ cells, GATA3 and FoxA1 decreased expression and ERα promoter regions became methylated, consistent with the loss of ERα expression. Lung metastases consisted of mostly ERα−RFP+Erbb2+ cells, a few ERα−RFP−Erbb2+ cells, and no ERα+RFP+Erbb2+ cells. The high metastatic capacity of ERα−RFP+Erbb2+ cells was associated with ERK1/2 activation. These results show that the slow-proliferating, nonmetastatic ERα+RFP+Erbb2+ cells progressively lose ERα during tumorigenesis to become fast-proliferating, highly metastatic ERα−RFP+Erbb2+ cells. The ERα−Erbb2+ BrCs with an ERα+ origin are more aggressive than those ERα−Erbb2+ BrCs with an ERα− origin, and thus, they should be distinguished and treated differently in the future.

How Lineage Tracing Studies Can Unveil Tumor Heterogeneity in Breast Cancer

“…Similar to the aforementioned studies, the activation of certain oncogenes in the luminal compartment was sufficient to develop tumors with either luminal, basal, or mixed (Keratin-8 and Keratin-5-positive) features [90]. [48,66], claudin-low (or mesenchymal-like) [96], and basal-like [18,73] tumors. Notes: this figure only includes studies that have used comparisons with human breast cancer transcriptomic datasets (PAM50, Hu306 or similar); studies performed with non-specific Cre lines, such as MMTV-Cre or Krt8-Cre ERT2 (which labels all MECs, or both LC populations, respectively), were discarded.…”

“…The combination of multiple in vivo systems (Cre/lox, RCAS/TVA, Dre/rox, and Flp/Frt) could be the key to designing a model able to recapitulate the tumor evolution observed in human breast cancer. [48,66], claudin-low (or mesenchymal-like) [96], and basal-like [18,73] tumors. Notes: this figure only includes studies that have used comparisons with human breast cancer transcriptomic datasets (PAM50, Hu306 or similar); studies performed with non-specific Cre lines, such as MMTV-Cre or Krt8-Cre ERT2 (which labels all MECs, or both LC populations, respectively), were discarded.…”

“…Fascinatingly, a recent work combined lineage tracing analysis and the RCAS-TVA system. This consists of expressing the TVA cell surface receptor under the control of a specific target promoter recognized by an avian leukosis virus-derived vector (RCAS) [ 47 ] to elucidate the contribution of ERα during cancer progression and metastasis of HER2-positive tumors [ 48 ]. By using Esr1 -Cre/ MMTV -TVA/ Rosa 26-tdRFP mice infected with RCAS- Erbb2 (to generate HER2-positive tumors), the authors could demonstrate that ERα pos tumor cells with an overexpression of HER2 have to progressively lose their ERα expression in order to clonally expand and metastasize [ 48 ].…”

“…This consists of expressing the TVA cell surface receptor under the control of a specific target promoter recognized by an avian leukosis virus-derived vector (RCAS) [ 47 ] to elucidate the contribution of ERα during cancer progression and metastasis of HER2-positive tumors [ 48 ]. By using Esr1 -Cre/ MMTV -TVA/ Rosa 26-tdRFP mice infected with RCAS- Erbb2 (to generate HER2-positive tumors), the authors could demonstrate that ERα pos tumor cells with an overexpression of HER2 have to progressively lose their ERα expression in order to clonally expand and metastasize [ 48 ]. Importantly, HER2-positive cells originated from ERα pos cells were more aggressive than those originated from ERα neg cells, suggesting that the cell of origin plays an important role in the clinical outcome of breast cancer patients.…”

“… Schematic representation of the mammary cellular plasticity during tumorigenesis. The use of different murine Cre lines ( Lgr5 -Cre ERT2 , Krt5 -Cre ERT2 or Krt14 -Cre to target BCs; Bgl -Cre or Wap -Cre to trace ERα neg LCs; and Esr1 -Cre to monitor ERα pos LCs) to induce the expression of specific mutations in different mammary epithelial cell types resulted in the generation of tumors that resembled some of the human subtypes: luminal A [ 43 ], luminal B [ 43 , 89 ], HER2-enriched [ 48 , 66 ], claudin-low (or mesenchymal-like) [ 96 ], and basal-like [ 18 , 73 ] tumors. Notes: this figure only includes studies that have used comparisons with human breast cancer transcriptomic datasets (PAM50, Hu306 or similar); studies performed with non-specific Cre lines, such as MMTV -Cre or Krt8 -Cre ERT2 (which labels all MECs, or both LC populations, respectively), were discarded.…”