Genetic Construction and Molecular Characterization of Breast Cancer Precursor Cells.
Abstract
Mutations in genes involved in cell-cycle and growth regulation, such as the retinoblastoma gene (RB), are expected to have a significant role in the early stages of breast cancer. RB-deficient cells are being created by two distinct methods. First, targeting vectors, currently being constructed, will be transfected into human mammary epithelial cell lines (HMEC) in order to knock out both RB allels via homologous recombination. Second, a human papilloma virus (HPV) E6/E7 fusion construct, previously shown to specifically target the retinoblastoma protein (pRb) for degradation, will be transfected into HMEC. The E6/E7 gene fusion will be introduced into the HMEC under the control of a regulatable promoter, allowing for direct regulation of protein expression. Breast cells made deficient for pRb by these two methods will be examined for cellular and molecular alterations, such as changes in morphology and behavior, and their ability to form duct-like structures when grown on or within an extracellular matrix. Additionally, since the normal RB product has been implicated in transcriptional regulation, the effect on mRNA transcription in the RB-deficient breast cells will be assayed by Northern blot analysis and differential display.
Document Details
- Document Type
- Technical Report
- Publication Date
- Jun 30, 1995
- Accession Number
- ADA298514
Entities
People
- Kimberly M. Spancake
Organizations
- University of Utah