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Deletion of PTEN Promotes Tumorigenic Signaling, Resistance to Anoikis, and Altered Response to Chemotherapeutic Agents in Human Mammary Epithelial Cells

¹ University of Maryland Greenebaum Cancer Center; ² University of Maryland Graduate Program in Molecular Medicine; Departments of ³ Biochemistry and Molecular Biology and ⁴ Radiation Oncology, University of Maryland School of Medicine; ⁵ The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Department of Oncology, The Johns Hopkins University School of Medicine, Baltimore, Maryland; and ⁶ Lombardi Comprehensive Cancer Center, Georgetown University School of Medicine, Washington, District of Columbia

Requests for reprints: Kurtis E. Bachman, GlaxoSmithKline, 709 Swedeland Road, UW2109, King of Prussia, PA 19406. Phone: 610-270-6045; Fax: 610-270-5598; E-mail: kurtis.e.bachman{at}gsk.com; Michele I. Vitolo, University of Maryland Greenebaum Cancer Center, 655 West Baltimore Street, Baltimore, MD 21201. Phone: 410-706-4142; Fax: 410-706-7619; E-mail: mvitolo{at}umaryland.edu; and David J. Weber, University of Maryland School of Medicine, 108 North Greene Street, Baltimore, MD 21201. Phone: 410-706-4354; Fax: 410-706-0458; E-mail: dweber{at}umaryland.edu.

Many cancers, including breast cancer, harbor loss-of-function mutations in the catalytic domain of phosphatase and tensin homologue deleted on chromosome 10 (PTEN) or have reduced PTEN expression through loss of heterozygosity and/or epigenetic silencing mechanisms. However, specific phenotypic effects of PTEN inactivation in human cancer cells remain poorly defined without a direct causal connection between the loss of PTEN function and the development or progression of cancer. To evaluate the biological and clinical relevance of reduced or deleted PTEN expression, a novel in vitro model system was generated using human somatic cell knockout technologies. Targeted homologous recombination allowed for a single and double allelic deletion, which resulted in reduced and deleted PTEN expression, respectively. We determined that heterozygous loss of PTEN in the nontumorigenic human mammary epithelial cell line MCF-10A was sufficient for activation of the phosphoinositide 3-kinase/AKT and mitogen-activated protein kinase pathways, whereas the homozygous absence of PTEN expression led to a further increased activation of both pathways. The deletion of PTEN was able to confer growth factor–independent proliferation, which was confirmed by the resistance of the PTEN^–/– MCF-10A cells to small-molecule inhibitors of the epidermal growth factor receptor. However, neither heterozygous nor homozygous loss of PTEN expression was sufficient to promote anchorage-independent growth, but the loss of PTEN did confer apoptotic resistance to cell rounding and matrix detachment. Finally, MCF-10A cells with the reduction or loss of PTEN showed increased susceptibility to the chemotherapeutic drug doxorubicin but not paclitaxel. [Cancer Res 2009;69(21):8275–83]

Key Words: PTEN • breast cancer • anoikis • tumor dormancy • doxorubicin