Mutant Breast Cancer Cell Lines Research Articles

Phosphatidylinositol-3-OH Kinase or RAS Pathway Mutations in Human Breast Cancer Cell Lines

Constitutive activation of the phosphatidylinositol-3-OH kinase (PI3K) and RAS signaling pathways are important events in tumor formation. This is illustrated by the frequent genetic alteration of several key players from these pathways in a wide variety of human cancers. Here, we report a detailed sequence analysis of the PTEN, PIK3CA, KRAS, HRAS, NRAS, and BRAF genes in a collection of 40 human breast cancer cell lines. We identified a surprisingly large proportion of cell lines with mutations in the PI3K or RAS pathways (54% and 25%, respectively), with mutants for each of the six genes. The PIK3CA, KRAS, and BRAF mutation spectra of the breast cancer cell lines were similar to those of colorectal cancers. Unlike in colorectal cancers, however, mutational activation of the PI3K pathway was mutually exclusive with mutational activation of the RAS pathway in all but 1 of 30 mutant breast cancer cell lines (P = 0.001). These results suggest that there is a fine distinction between the signaling activators and downstream effectors of the oncogenic PI3K and RAS pathways in breast epithelium and those in other tissues.

Developmental Profiles of Rb and Ras in Breast Cancer

Objective: Both Ras and retinoblastoma (Rb) play a significant role in cell proliferation and various differentiation programs in human reproduction. It has been reported that Rb-deficient, but not Rb-positive fibroblasts showed the elevation of activated N- and K-ras. In order to investigate the developmental process of breast cancer and to diagnose the early stage of breast cancer, we screened the expression of oncogenes including Rb (tumor suppressor) and Ras. Design: A variety of cell lines for breast cancers (MCF7, T-47D, MDA-MB-468, and BT-549) had been collected and tested for the sequence and expression of different oncogenes. Materials and Methods: Breast cancer cells were culture in DMEM medium supplemented with 10% FBS, sequence of Rb gene in different breast cancer cells was determined, and the protein expression of this gene was tested. And the Activated Ras Interaction Assay (ARIA) was carried out by GST-pull down followed by western blotting using anti-Ras antibodies. Activated Ras-GTP form was precipitated with the Ras binding domain of Raf-1 immobilized by fusion to GST and binding to glutathione beads. Results: In a normal Rb-expressing breast cancer cell line MCF-7, the Rb gene is genetically normal, and either the activity of N-ras or H-ras is increased. However, in a mutant Rb-expressing breast cancer cell line MDA-MB-468, the Rb gene is genetically abnormal, and the activity of both K-ras and N-ras is increased. legendCell lineslegendND = not determined.Rb geneRb expressionRas expressionMCF-7NormalNormalIncreased (N or H?)T-47DNormalNormalNDMDA-MB-468MutantNo expressionIncreased (N & K)BT-549MutantNo expressionNDlegend ND = not determined. Open table in a new tab Conclusion: Even though they are all breast cancer cell lines, the expression of ras proteins is different based on the expression of Rb genes. Our results indicate that the expression of 3 different ras proteins (H-, K-, and N-ras) has a relationship with the genetic constitution of Rb. Therefore, this developmental profile for both Ras and Rb would be helpful in identifying the early stage of breast cancer.