What changes in cellular function account for the progression of breast cancer? Onepromising research area that addresses this fundamental question is the study of the interactions of insulin-like growth factor-II (IGF-II) and catbepsin D with the IGF-II/mannose-6-phosphale (M6P) receptor. IGF-II is a mitogen for breast cancer and stimulates cell motility important for metastasis. Cathepsin D is an enzyme involved in metastasis. Both bind the IGF-II/M6P receptor at distinct binding sites. Nevertheless, a reciprocal inhibitory interaction is observed when IGF-II or catbepsin D bind the IGF-II/M6P receptor. Normally, cathepsin D is stored intracellularly in the lysosomes. However, a significant amount of this protease is secreted in breast cancer. We have developed an in vitro model consisting of MCF-7 breast cancer cells stably transfected with IGF-II. The expression of IGF-II in these transfected cells significantly increases the secretion of cathepsin D. Since higher levels of IGF-II and cathepsin D represent an increased risk of malignancy and metastasis, we hypothesize that inhibition of IGF-II expression will decrease cathepsin D, thus decreasing malignancy and metastatic potential. To address this question, MCF-7 breast cancer cells overexpressing IGF-II were transfected with an antisense IGF-II construct Stable transfectants were cloned by limiting dilution in 96 well plates. Clones with reduced or no IGF-II were identified by performing a dot blot assay on aliquots of serum-free conditioned media Western blotting analysis to characterize the IGF-II forms on antisense transfected clones demonstrated that these clones expressed only the 14 kD IGF-II form. In contrast, IGF-II secreting clones expressed several forms of IGF-II ranging from 14-45 kD. The total expression of IGF-II in antisense clones was reduced approximately by ten fold when compared to IGF-II secreting clones. Northern blotting analysis revealed that expression of the IGF-II mRNA was also significantly decreased by approximately five fold in antisense clones. Preliminary cathepsin D analysis by western blotting showed that antisense IGF-II clones secreted less cathepsin D. Since IGF-II and cathepsin D promote tumor growth and metastasis, inhibition of these proteins represents a desirable target for treatment. Thus, regulation of IGF-II may offer a unique therapeutic target for a molecular medicine approach to the treatment of breast cancer.
|Original language||English (US)|
|Journal||Journal of Investigative Medicine|
|State||Published - Jan 1 1996|
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)