Human breast cancer cells generated by oncogenic transformation of primary mammary epithelial cells

Elenbaas, Brian; Spirio, Lisa; Koerner, Frederick C.; Fleming, Mark D.; Zimonjic, Drazen B.; Donaher, Joana Liu; Popescu, Nicholas C.; Hahn, William C.; Weinberg, Robert A.

doi:10.1101/gad.828901

Cited by 793 publications

(887 citation statements)

References 46 publications

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“…In BJ cells, oncogenic RAS expression upregulates MTH1 levels ( Figure 1a). Similarly, when we compared MTH1 expression levels in MCF7 human breast cancer cells versus their transformed derivatives, MCF7-RAS (Kasid et al, 1985), as well as in HMLE versus HMLE-RAS cells (experimentally immortalized and transformed human mammary epithelial cells) (Elenbaas et al, 2001), we found that in each case MTH1 levels were higher in the oncogenic RAS-transformed counterpart cells (Figure 4a). Thus, we undertook to determine whether suppression of MTH1 expression would affect the proliferation of RAS-transformed cells relative to the corresponding non-oncogenic RAS-expressing cell line.…”

Section: Resultsmentioning

confidence: 81%

“…MCF7 and MCF7-RAS cell lines were maintained in DMEM supplemented with 10% fetal calf serum, 100 units/ml penicillin, 100 mg/ml streptomycin and 2 mM L-glutamine at 37 1C in either 21% oxygen/5% CO 2 or, where specified, in 3% oxygen/5% CO 2 . HMLE and HMLE-RAS breast epithelial cell lines were derived from human mammary epithelial cells (HMECs) from Clonetics and maintained in supplemented MEGM media (Clonetics/Lonza, Walkersville, MD, USA), as described (Elenbaas et al, 2001). The shRNA design, lentivirus production and infection was done as described (Stewart et al, 2003;Rai et al, 2009).…”

Section: Resultsmentioning

confidence: 99%

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Enhanced elimination of oxidized guanine nucleotides inhibits oncogenic RAS-induced DNA damage and premature senescence

et al. 2010

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Approximately 20% of tumors contain activating mutations in the RAS family of oncogenes. As tumors progress to higher grades of malignancy, the expression of oncogenic RAS has been reported to increase, leading to an oncogene-induced senescence (OIS) response. Evasion of this senescence barrier is a hallmark of advanced tumors indicating that OIS serves a critical tumorsuppressive function. Induction of OIS has been attributed to either RAS-mediated production of reactive oxygen species (ROS) or to induction of a DNA damage response (DDR). However, functional links between these two processes in triggering the senescent phenotype have not been explicitly described. Our previous work has shown that, in cultured untransformed cells, preventing elimination of oxidized guanine deoxyribonucleotides, which was achieved by suppressing expression of the cellular 8-oxodGTPase, human MutT homolog 1 (MTH1), sufficed to induce a DDR as well as premature senescence. Here, we demonstrate that overexpression of MTH1 can prevent the oncogenic H-RAS-induced DDR and attendant premature senescence, although it does not affect the observed elevation in ROS levels produced by RAS oncoprotein expression. Conversely, we find that loss of MTH1 preferentially induces an in vitro proliferation defect in tumorigenic cells overexpressing oncogenic RAS. These results indicate that the guanine nucleotide pool is a critical target for intracellular ROS produced by oncogenic RAS and that RAS-transformed cells require robust MTH1 expression to proliferate.

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Section: Resultsmentioning

confidence: 81%

Section: Resultsmentioning

confidence: 99%

Enhanced elimination of oxidized guanine nucleotides inhibits oncogenic RAS-induced DNA damage and premature senescence

et al. 2010

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“…To explore whether ferroportin is present in normal human breast epithelial cells and whether its concentrations are altered in breast cancer, we compared ferroportin protein abundance in three pairs of mammary epithelial cell types with variable malignant potential: (i) primary normal human mammary epithelial (HME) cells and tumor-forming variants of these cells derived by sequential transformation of HME cells with the catalytic subunit of telomerase, SV40 T antigen, and high levels of oncogenic H- ras (25) (termed R5 cells here); (ii) MCF10A cells, a spontaneously immortalized diploid cell line obtained from reduction mammoplasty (26), and MCF7 (27), a breast cancer cell line established from a pleural effusion in a patient with metastatic breast cancer; and (iii) SUM102 cells, a breast epithelial cell line with a normal karyotype isolated from early-stage breast cancer (28), and SUM149, a cell line developed from an aggressive inflammatory breast cancer (29). Examination of ferroportin in these cells revealed that protein abundance was reduced in all aggressive breast cancer cell lines when compared to their counterparts with little or no malignant potential (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…HME cells transduced with h- TERT , SV40 T antigen, and high levels of H- ras are termed R5 cells here and were a gift from the laboratory of R. Weinberg (25). All cells were maintained at 37°C in a humidified atmosphere containing 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

Ferroportin and Iron Regulation in Breast Cancer Progression and Prognosis

Pinnix¹,

Miller²,

Wang³

et al. 2010

Science Translational Medicine

245

278

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Ferroportin and hepcidin are critical proteins for the regulation of systemic iron homeostasis. Ferroportin is the only known mechanism for export of intracellular non–heme-associated iron; its stability is regulated by the hormone hepcidin. Although ferroportin profoundly affects concentrations of intracellular iron in tissues important for systemic iron absorption and trafficking, ferroportin concentrations in breast cancer and their influence on growth and prognosis have not been examined. We demonstrate here that both ferroportin and hepcidin are expressed in cultured human breast epithelial cells and that hepcidin regulates ferroportin in these cells. Further, ferroportin protein is substantially reduced in breast cancer cells compared to nonmalignant breast epithelial cells; ferroportin protein abundance correlates with metabolically available iron. Ferroportin protein is also present in normal human mammary tissue and markedly decreased in breast cancer tissue, with the highest degree of anaplasia associated with lowest ferroportin expression. Transfection of breast cancer cells with ferroportin significantly reduces their growth after orthotopic implantation in the mouse mammary fat pad. Gene expression profiles in breast cancers from >800 women reveal that decreased ferroportin gene expression is associated with a significant reduction in metastasis-free and disease-specific survival that is independent of other breast cancer risk factors. High ferroportin and low hepcidin gene expression identifies an extremely favorable cohort of breast cancer patients who have a 10-year survival of >90%. Ferroportin is a pivotal protein in breast biology and a strong and independent predictor of prognosis in breast cancer.

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“…For example, introduction of HPV E6 and E7 into primary HMECs led to their immortalization (Wazer et al, 1995;Gudjonsson et al, 2002). In another study, introduction of Simian Virus 40 Early Region (SV40ER) and the catalytic subunit of human telomerase (hTERT) into adherent HMECs led to the generation of immortalized HMLE cells (Elenbaas et al, 2001). More recently, introduction of hTERT-alone was shown to suffice for the immortalization of HMECs grown in a specialized media (Zhao et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

Introduction of SV40ER and hTERT into mammospheres generates breast cancer cells with stem cell properties

et al. 2011

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Human breast cancer cells generated by oncogenic transformation of primary mammary epithelial cells

Cited by 793 publications

References 46 publications

Enhanced elimination of oxidized guanine nucleotides inhibits oncogenic RAS-induced DNA damage and premature senescence

Enhanced elimination of oxidized guanine nucleotides inhibits oncogenic RAS-induced DNA damage and premature senescence

Ferroportin and Iron Regulation in Breast Cancer Progression and Prognosis

Introduction of SV40ER and hTERT into mammospheres generates breast cancer cells with stem cell properties

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