This proposal will study non-MDR1 genetic and cellular determinants of taxane therapy in breast and ovarian cancers. We have established 17 taxane-resistant variants from 4 breast and 6 ovarian cancer cell lines, and we hypothesize that these diverse variants may yield insight into novel mechanisms of resistance to taxanes.
The specific aims are: 1. To determine tubulin isoform content and potential alterations in microtubule dynamic instability. Our analyses of the expression of the six classes of (3- tubulin isoforms reveal altered p-tubulin content in many of the taxane-resistant variants. Microtubule assembly in these cells will be assessed in collaboration with Dr. Mary Ann Jordan. 2. To study the effects of altered tubulin and microtubule associated protein genes on drug sensitivity. Several of the variants have down-regulated MAP4 and up-regulated MAP Tau, and we hypothesize that these alterations may have an adverse effect on taxane sensitivity by altering taxane binding. We plan to use siRNA technology to study the effects of these genes and specific tubulin isoforms, and inducible vectors utilizing tetracycline regulation will be used to transfect genes of interest into parental cells. 3. To explore regulators of apoptosis (e.g. Bcl-2, Bcl-XL, Akt) and cell growth (e.g. EGFR-1, her2) as determinants of response to taxanes. Several of the non-P-gp variants possess altered expression of apoptotic regulators. We will use antisense and RNAi approaches to study the effects of inhibition of these regulators on cellular sensitivity to taxanes. We will also determine the effects of inhibition of growth pathways on taxane sensitivity by inhibiting EGFR-1, her2, Akt and other genes using kinase inhibitors. 4. To identify novel genetic markers to predict resistance to taxanes. Genetic (array CGH) and genomic (microarray gene expression profiling) approaches will be used to search for novel genetic markers for the prediction of sensitivity and resistance to taxane therapy. Candidate genes will be validated by PCR, immunoblotting, and tet-regulated expression using an MCF-7 Flp-ln line which we have made. 5. To study taxane resistance in clinical specimens of breast and ovarian cancer. Whole genome profiling of tumor specimens from breast and ovarian cancers from the NCI Cooperative Human Tissue Network, annotated for responsiveness to taxane therapy, will be performed. Expression of resistance factors identified in cellular models will also be validated in these specimens. In summary, this project will use a large set of unique models of cellular resistance to taxanes in ovarian and breast cancer cell lines and clinical specimens obtained from patients treated with taxanes to: (1) study the relevance of known resistance mechanisms to these drugs, (2) explore new mechanisms, and (3) develop clinical markers for response.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA114037-04
Application #
7826596
Study Section
Drug Discovery and Molecular Pharmacology Study Section (DMP)
Program Officer
Forry, Suzanne L
Project Start
2007-08-01
Project End
2012-05-31
Budget Start
2010-06-01
Budget End
2011-05-31
Support Year
4
Fiscal Year
2010
Total Cost
$297,730
Indirect Cost
Name
Stanford University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94305
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Duran, George E; Wang, Yan C; Moisan, François et al. (2017) Decreased levels of baseline and drug-induced tubulin polymerisation are hallmarks of resistance to taxanes in ovarian cancer cells and are associated with epithelial-to-mesenchymal transition. Br J Cancer 116:1318-1328
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