Suppressor Genes in Breast Cancer.
Annual rept. 1 Jul 97-30 Jun 98
GEORGETOWN UNIV WASHINGTON DC
Pagination or Media Count:
The inactivation of tumor suppressor genes TSGs in a variety of human cancers has inspired an intense search for the breast cancer-specific TSGs. The purposes of this proposal are to 1 clone novel TSGs specific to human breast cancer 2 examine the alteration of these TSGs in primary breast-tumors and ultimately 3 identify their characteristics, regulation and function. We have constructed a cDNA library from normal human breast epithelia and cloned it into a vector that is negatively regulated by the tet repressor tetR and simultaneously expresses the enhanced green fluorescent protein. These vectors were then co-transfected into MCF-7 cells that express tetR. Upon withdrawal of tet, the repressed expression of the cDNA of interest is released and the cDNA is expressed. Using a novel dye that is retained in nonproliferating cells, we were able to identify growth inhibited clones. These clones were then sorted by Flow Cytometry. This functional screen has provided the basis for cloning TSGs which are expressed in the growth inhibited cells. Using PCR, we can obtain the insert sequence and then screen primary breast carcinomas to discover muntations in cloned TSGs.
- Anatomy and Physiology
- Medicine and Medical Research