The direct effect of focal adhesion kinase (FAK), dominant-negative FAK, FAK-CD and FAK siRNA on gene expression and human MCF-7 breast cancer cell tumorigenesis.

Abstract

BACKGROUND:

Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase that plays an important role in survival signaling. FAK has been shown to be overexpressed in breast cancer tumors at early stages of tumorigenesis.

METHODS:

To study the direct effect of FAK on breast tumorigenesis, we developed Tet-ON (tetracycline-inducible) system of MCF-7 breast cancer cells stably transfected with FAK or dominant-negative, C-terminal domain of FAK (FAK-CD), and also FAKsiRNA with silenced FAK MCF-7 stable cell line. Increased expression of FAK in isogenic Tet-inducible MCF-7 cells caused increased cell growth, adhesion and soft agar colony formation in vitro, while expression of dominant-negative FAK inhibitor caused inhibition of these cellular processes. To study the role of induced FAK and FAK-CD in vivo, we inoculated these Tet-inducible cells in nude mice to generate tumors in the presence or absence of doxycycline in the drinking water. FAKsiRNA-MCF-7 cells were also injected into nude mice to generate xenograft tumors.

RESULTS:

Induction of FAK resulted in significant increased tumorigenesis, while induced FAK-CD resulted in decreased tumorigenesis. Taq Man Low Density Array assay demonstrated specific induction of FAKmRNA in MCF-7-Tet-ON-FAK cells. DMP1, encoding cyclin D binding myb-like protein 1 was one of the genes specifically affected by Tet-inducible FAK or FAK-CD in breast xenograft tumors. In addition, silencing of FAK in MCF-7 cells with FAK siRNA caused increased cell rounding, decreased cell viability in vitro and inhibited tumorigenesis in vivo. Importantly, Affymetrix microarray gene profiling analysis using Human Genome U133A GeneChips revealed >4300 genes, known to be involved in apoptosis, cell cycle, and adhesion that were significantly down- or up-regulated (p < 0.05) by FAKsiRNA.

CONCLUSION:

Thus, these data for the first time demonstrate the direct effect of FAK expression and function on MCF-7 breast cancer tumorigenesis in vivo and reveal specific expression of genes affected by silencing of FAK.

• Authors: Vita M Golubovskaya, Min Zheng, Li Zhang, Jian-Liang Li, William G Cance
• Journal: BMC cancer (BMC Cancer) Vol. 9 Pg. 280 (Aug 12 2009) ISSN: 1471-2407 [Electronic] England
• PMID: 19671193 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• Antineoplastic Agents
• RNA, Small Interfering
• Focal Adhesion Kinase 1
• Doxycycline

Topics

• Animals
• Antineoplastic Agents (pharmacology)
• Breast Neoplasms (metabolism)
• Cell Line, Tumor
• Doxycycline (pharmacology)
• Female
• Focal Adhesion Kinase 1 (genetics, metabolism)
• Genome, Human
• Humans
• Mice
• Mice, Nude
• Neoplasm Transplantation
• Oligonucleotide Array Sequence Analysis
• RNA, Small Interfering (metabolism)
• Transfection