There is increasing evidence that breast and other cancers originate from and are maintained by a small fraction of stem/progenitor cells with self-renewal properties. Whether such cancer stem/progenitor cells originate from normal stem cells based on initiation of a de novo stem cell program, by reprogramming of a more differentiated cell type by oncogenic insults, or both remains unresolved. A major hurdle in addressing these issues is lack of immortal human stem/progenitor cells that can be deliberately manipulated in vitro. We present evidence that normal and human telomerase reverse transcriptase (hTERT)-immortalized human mammary epithelial cells (hMECs) isolated and maintained in Dana-Farber Cancer Institute 1 (DFCI-1) medium retain a fraction with progenitor cell properties. These cells coexpress basal (K5, K14, and vimentin), luminal (E-cadherin, K8, K18, or K19), and stem/progenitor (CD49f, CD29, CD44, and p63) cell markers. Clonal derivatives of progenitors coexpressing these markers fall into two distinct types-a K5 + /K19− type and a K5 + / K19 + type. We show that both types of progenitor cells have selfrenewal and differentiation ability. Microarray analyses confirmed the differential expression of components of stem/progenitorassociated pathways, such as Notch, Wnt, Hedgehog, and LIF, in progenitor cells compared with differentiated cells. Given the emerging evidence that stem/progenitor cells serve as precursors for cancers, these cellular reagents represent a timely and invaluable resource to explore unresolved questions related to stem/progenitor origin of breast cancer.reast cancer is a genetically and clinically heterogeneous disease (1). It is unclear whether different target cells contribute to this heterogeneity and which cell types are most susceptible to oncogenesis. Recent molecular profiling has identified five major subtypes of breast cancers: a basal epithelium-like group, an ErbB2-overexpressing group, a normal breast epithelium-like group, and two luminal epithelial cell types with significantly different outcomes for patients belonging to various groups (2). The correspondence of some breast cancer subtypes with cell types present in the normal mammary gland (such as basal and luminal) strongly supports the idea that breast tumor subtypes may represent malignancies of biologically distinct cell subtypes. Alternatively, different types of breast cancers may arise from a common precursor, based on distinct pathways of oncogene-driven reprogramming. Heterogeneity in cancers is ascribed to clonal evolution as a result of inherent genomic instability of tumor cells and tumor-host interactions (3). The stem cell hypothesis suggests an alternate explanation with tumor heterogeneity reflecting the relative fraction of cancer stem/progenitor cells and differences in their abilities to produce progeny at various stages of differentiation. Although current literature supports each of these ideas, definitive studies to favor one or the other model, or both hypotheses, are lacking.Recent prog...

Background-Our objective was to determine whether abciximab, eptifibatide, or tirofiban inhibited ligand binding to ␣ v ␤ 3 integrins on human aortic smooth muscle cells (HASMCs) or human umbilical vein endothelial cells (HUVECs). Abciximab binds ␣ IIb ␤ 3 on platelets and ␣ v ␤ 3 on HUVECs with similar affinity, whereas eptifibatide and tirofiban are thought to be highly specific for ␣ IIb ␤ 3 . The conclusion that eptifibatide does not bind vascular ␣ v ␤ 3 integrins may be premature, however, because recent studies have demonstrated that the affinity of ␣ v ␤ 3 for various ligands, including antagonists, is subject to modulation. Methods and Results-Abciximab and 7E3, the anti-␤ 3 integrin monoclonal antibody from which abciximab was derived, bound ␣ v ␤ 3 on HASMCs in a specific and saturable manner and with an affinity similar to binding to ␣ IIb ␤ 3 on platelets. 7E3 and eptifibatide inhibited ␣ v ␤ 3 -mediated attachment of HASMCs to thrombospondin (TSP) and prothrombin but had no effect on ␣ v ␤ 5 -or ␤ 1 -mediated HASMC attachment to vitronectin-, collagen-, or fibronectin-coated or noncoated tissue culture plates. The inhibitory effect of eptifibatide was similar in magnitude and not additive to that of 7E3.

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Transforming growth factor (TGF)-beta1 has been implicated in vascular healing responses after mechanical injury. Using cultured rat aortic smooth muscle cells (RASMC), we examined the hypothesis that production and secretion of thrombospondin (TSP) contributes to TGF-beta1-induced proliferation. We found that TGF-beta1 enhanced production and secretion of TSP, with peak levels of secreted TSP observed 24 h after treatment. RASMC treated with TGF-beta1 secreted a mitogenic activity that was transferable in conditioned media and partially inhibited by C6.7, a monoclonal anti-TSP antibody. Exogenous TSP stimulated a proliferative response, with maximal [(3)H]thymidine incorporation occurring 24 h earlier than maximal [(3)H]thymidine incorporation in response to TGF-beta1-treatment. Pretreatment with C6.7 or polyclonal anti-TSP neutralizing antibodies inhibited TGF-beta1-induced proliferation of RASMC. Proliferative responses to TGF-beta1 were also inhibited by pretreatment with an anti-beta(3) integrin monoclonal blocking antibody (F11), RGD peptides, and the anti-alpha(v)beta(3) disintegrin echistatin. Treatment with TSP and TGF-beta1 increased c-Jun NH(2)-terminal kinase (JNK)1 activity, with peak effects observed at 15 min and 4 h, respectively. Treatment with C6.7 or F11 inhibited TGF-beta-induced activation of JNK1. In summary, these studies support the hypothesis that TGF-beta-induced JNK1 activation and proliferation of RASMC require secretion of TSP and ligation of alpha(v)beta(3)-integrins.