Alternate Splicing of CD44 Messenger RNA in Prostate Cancer Growth
Abstract
Aim 1: Loss of CD44 standard and increased splice variant form CD44v7-10 facilitate prostate cancer (PC) invasion. First Sub-Aim: A manuscript was published (attached) on the role of Mitogen-activated protein kinase (MAPK) pathways and paracrine calcitonin, both of which dysregulate CD44. Second Sub-Aim: Metabolic labeling studies of CD44 total and CD44v7-10 protein were pursued over about a 6- month period, but the findings were not publishable. Aim 2: The use of adeno-associated virus for altering expression of CD44 prior to in vitro and in vivo studies was prohibitive owing to viral cytopathic effect. Instead, we used lentiviral or retroviral transfection methods in PC-3M cells, for stably altered expression. Confirmation of re-expression of CD44s as a Fusion protein (with luciferase) or Separate protein, or of RNAi knockdown of CD44v7-10, was achieved using qRT-PCR, western blot analysis, and IVIS visualization of luminescence after adding luciferin substrate, in a flask or mouse tumor. Cells re-expressing CD44s as a Separate protein had decreased growth, decreased Matrigel migration and invasion, decreased anchorage independent colony formation, restoration of adhesion to hyaluronan (a benign feature), and apparently less tumor take and growth rate of subcutaneous xenografts in mice. Whether CD44s re-expression influences chemosensitivity to Docetaxel will require more experiments. Expression of CD44s as a Fusion protein was less definitive in assays. Effects of RNAi to CD44v7-10 are not clear yet.
Document Details
- Document Type
- Technical Report
- Publication Date
- Apr 01, 2009
- Accession Number
- ADA506471
Entities
People
- Kenneth A. Iczkowski
Organizations
- University of Colorado Health