Access Type
Open Access Dissertation
Date of Award
January 2011
Degree Type
Dissertation
Degree Name
Ph.D.
Department
Cancer Biology
First Advisor
Bonnie F. Sloane
Abstract
FUNCTIONAL IN VITRO ANALYSES OF LIPID RAFT-ASSOCIATED CATHEPSIN B: IMPLICATION FOR THE INVASIVE PHENOTYPE
OF INFLAMMATORY BREAST CANCER
by
BERNADETTE C. VICTOR
December 2011
Advisor: Dr. Bonnie F. Sloane
Major: Cancer Biology
Degree: Doctor of Philosophy
Inflammatory breast cancer (IBC) is an aggressive, metastatic and
highly angiogenic form of locally advanced breast cancer. Breast
cancer invasion has been linked to proteolytic activity at the tumor
cell surface. We observed that uPA, uPAR and enzymatically active
cathepsin B were all present in caveolae fractions isolated from
SUM149 cells. Using a live-cell proteolysis assay, we demonstrated
that both IBC cell lines degrade type IV collagen. The SUM149 cells
exhibit predominantly pericellular proteolysis, consistent with
localization of constituents of a proteolytic pathway to membrane
microdomains. A functional role for cathepsin B was confirmed by
the ability of CA074, a cell impermeable and highly selective
cathepsin B inhibitor, to significantly reduce pericellular proteolysis
and invasion by SUM149 cells. A statistically significant co-
expression of cathepsin B and caveolin-1 was found in IBC patient
biopsies, thus validating our in vitro data. To determine whether the
uPA and cathepsin B present at the cell surface is localized
specifically to caveolae and investigate if caveolae are a functional
component of the IBC phenotype, we knocked down the expression
of caveolin-1 in SUM149 cells. The subcellular distribution of
cathepsin B was unchanged in caveolin-1 knockdown SUM149 cells.
In contrast, knockdown decreased the association of both uPA and
EGFR with caveolae. Knockdown of caveolin-1 also decreased
degradation of type IV collagen and invasion by IBC cells. We also
determined that targeted disruption of lipid microdomains by use of
lovastatin diminished the aggressiveness of IBC cells. Cathepsin B
activity and association with lipid microdomains was reduced in
lovastatin treated IBC cells. Lovastatin treatment also reduced type
IV collagen degradation and invasion by IBC cells. In conclusion, we
have shown that cathepsin B and caveolin-1 both contribute to the
aggressiveness of IBC, albeit by different mechanisms.
Recommended Citation
Victor, Bernadette Caroline, "Functional in vitro analyses of lipid raft-associated cathepsin b: implication for the invasive phenotype of inflammatory breast cancer" (2011). Wayne State University Dissertations. 400.
https://digitalcommons.wayne.edu/oa_dissertations/400