Dendritic cells (DCs) play a pivotal role in the tumor microenvironment, which is known to affect disease progression in many human malignancies. Infiltration by mature, active DCs into the tumors confers an increase in immune activation and recruitment of disease-fighting immune effector cells and pathways. DCs are the preferential target of infiltrating T cells. However, tumor cells have means of suppressing DC function or of altering the tumor microenvironment in such a way that immune-suppressive DCs are recruited. Advances in understanding these changes have led to promising developments in cancer-therapeutic strategies targeting tumor-infiltrating DCs to subdue their immunosuppressive functions and enhance their immune-stimulatory capacity.

The breast cancer stem cell (BCSC) hypotheses suggest that breast cancer is derived from a single tumor-initiating cell with stem-like properties, but the source of these cells is unclear. We previously observed that induction of an immune response against an epithelial breast cancer led in vivo to the T-cell–dependent outgrowth of a tumor, the cells of which had undergone epithelial to mesenchymal transition (EMT). The resulting mesenchymal tumor cells had a CD24^−/loCD44⁺ phenotype, consistent with BCSCs. In the present study, we found that EMT was induced by CD8 T cells and the resulting tumors had characteristics of BCSCs, including potent tumorigenicity, ability to reestablish an epithelial tumor, and enhanced resistance to drugs and radiation. In contrast to the hierarchal cancer stem cell hypothesis, which suggests that breast cancer arises from the transformation of a resident tissue stem cell, our results show that EMT can produce the BCSC phenotype. These findings have several important implications related to disease progression and relapse. [Cancer Res 2009;69(7):2887–95]

Despite significant progress in the treatment of breast cancer, particularly through the use of targeted therapy, relapse and chemoresistance remain a major hindrance to the fight to minimize the burden of the disease. It is becoming increasingly clear that a rare subpopulation of cells known as cancer stem cells (CSC), able to be generated through epithelial-to-mesenchymal transition (EMT) and capable of tumor initiation and self-renewal, contributes to treatment resistance and metastases. This means that a more effective therapy should target both the chemoresistant CSCs and the proliferating epithelial cells that give rise to them to reverse EMT and to attenuate their conversion to CSCs. Here, we demonstrate a novel function of AXL in acting upstream to induce EMT in normal and immortalized human mammary epithelial cells in an apparent positive feedback loop mechanism and regulate breast CSC (BCSC) self-renewal and chemoresistance. Downregulation of AXL using MP470 (Amuvatinib) reversed EMT in mesenchymal normal human mammary epithelial cells and murine BCSCs attenuating self-renewal and restored chemosensitivity of the BCSCs. AXL expression was also found to be associated with the expression of stem cell genes, regulation of metastases genes, increased tumorigenicity and was important for BCSC invasion and migration. Inactivation of AXL also led to the downregulation of nuclear factor-κB pathway and reduced tumor formation in vivo. Taken together, our data suggest that targeted therapy against AXL, in combination with systemic therapies, has the potential to improve response to anticancer therapies and to reduce breast cancer recurrence and metastases.

Breast cancer recurrence is believed to be caused by a subpopulation of cancer cells that possess the stem cell attribute of treatment resistance. Recently, we and others have reported the generation of breast cancer stem cells (BCSC) by epithelial–mesenchymal transition (EMT), although the physiologic process by which these cells may arise in vivo remains unclear. We show here that exposure of tumor cells to TGFβ and TNFα induces EMT and, more importantly, generates cells with a stable BCSC phenotype which is shown by increased self-renewing capacity, greatly increased tumorigenicity, and increased resistance to oxaliplatin, etoposide, and paclitaxel. Furthermore, gene expression analyses found that the TGFβ/TNFα-derived BCSCs showed downregulated expression of genes encoding claudin 3, 4, and 7 and the luminal marker, cytokeratin 18. These changes indicate a shift to the claudin-low molecular subtype, a recently identified breast cancer subtype characterized by the expression of mesenchymal and stem cell-associated markers and correlated with a poor prognosis. Taken together, the data show that cytokine exposure can be used to generate stable BCSCs ex vivo, and suggest that these cells may provide a valuable tool in the identification of stem cell-directed biomarkers and therapies in breast cancer. Cancer Res; 71(13); 4707–19. ©2011 AACR.

Within the ovarian cancer microenvironment, there are several mechanisms that suppress the actions of antitumor immune effectors. Delineating the complex immune microenvironment is an important goal toward developing effective immune-based therapies. A dominant pathway of immune suppression in ovarian cancer involves tumor-associated and dendritic cell (DC)-associated B7-H1. The interaction of B7-H1 with PD-1 on tumor-infiltrating T cells is a widely cited theory of immune suppression involving B7-H1 in ovarian cancer. Recent studies suggest that the B7-H1 ligand, programmed death receptor-1 (PD-1), is also expressed on myeloid cells, complicating interpretations of how B7-H1 regulates DC function in the tumor. In this study, we found that ovarian cancer-infiltrating DCs progressively expressed increased levels of PD-1 over time in addition to B7-H1. These dual-positive PD-1⁺ B7-H1⁺ DCs have a classical DC phenotype (i.e., CD11c⁺CD11b⁺CD8⁻), but are immature, suppressive, and respond poorly to danger signals. Accumulation of PD-1⁺B7-H1⁺ DCs in the tumor was associated with suppression of T cell activity and decreased infiltrating T cells in advancing tumors. T cell suppressor function of these DCs appeared to be mediated by T cell-associated PD-1. In contrast, ligation of PD-1 expressed on the tumor-associated DCs suppressed NF-κB activation, release of immune regulatory cytokines, and upregulation of costimulatory molecules. PD-1 blockade in mice bearing ovarian cancer substantially reduced tumor burden and increased effector Ag-specific T cell responses. Our results reveal a novel role of tumor infiltrating PD-1⁺B7-H1⁺ DCs in mediating immune suppression in ovarian cancer.