PTK6 Regulates IGF-1-Induced Anchorage-Independent Survival Hanna Y. Irie1,2, Yashaswi Shrestha2, Laura M. Selfors1, Fabianne Frye1, Naoko Iida1, Zhigang Wang 3 , Lihua Zou3, Jun Yao 2 , Yiling Lu4, Charles B. Epstein5,6, Sridaran Natesan 5 , Andrea L. Richardson 3 , Kornelia Polyak2, Gordon B. Mills4, William C. Hahn1,6, Joan S. Brugge1* 1 Department of Cell Biology, Harvard Medical School, Boston, Massachusetts, United States of America, 2 Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, United States of America, 3 Department of Pathology, Brigham and Women’s Hospital, Boston, Massachusetts, United States of America, 4 Department of Systems Biology, University of Texas MD Anderson Cancer Center, Houston, Texas, United States of America, 5 Sanofi-Aventis, Cambridge, Massachusetts, United States of America, 6 Broad Institute of Harvard and Massachusetts Institute of Technology (MIT), Cambridge, Massachusetts, United States of America Abstract Background: Proteins that are required for anchorage-independent survival of tumor cells represent attractive targets for therapeutic intervention since this property is believed to be critical for survival of tumor cells displaced from their natural niches. Anchorage-independent survival is induced by growth factor receptor hyperactivation in many cell types. We aimed to identify molecules that critically regulate IGF-1-induced anchorage-independent survival. Methods and Results: We conducted a high-throughput siRNA screen and identified PTK6 as a critical component of IGF-1 receptor (IGF-1R)-induced anchorage-independent survival of mammary epithelial cells. PTK6 downregulation induces apoptosis of breast and ovarian cancer cells deprived of matrix attachment, whereas its overexpression enhances survival. Reverse-phase protein arrays and subsequent analyses revealed that PTK6 forms a complex with IGF-1R and the adaptor protein IRS-1, and modulates anchorage-independent survival by regulating IGF-1R expression and phosphorylation. PTK6 is highly expressed not only in the previously reported Her2+ breast cancer subtype, but also in high grade ER+, Luminal B tumors and high expression is associated with adverse outcomes. Conclusions: These findings highlight PTK6 as a critical regulator of anchorage-independent survival of breast and ovarian tumor cells via modulation of IGF-1 receptor signaling, thus supporting PTK6 as a potential therapeutic target for multiple tumor types. The combined genomic and proteomic approaches in this report provide an effective strategy for identifying oncogenes and their mechanism of action. Citation: Irie HY, Shrestha Y, Selfors LM, Frye F, Iida N, et al. (2010) PTK6 Regulates IGF-1-Induced Anchorage-Independent Survival. PLoS ONE 5(7): e11729. doi:10.1371/journal.pone.0011729 Editor: Nils Cordes, Dresden University of Technology, Germany Received February 4, 2010; Accepted June 7, 2010; Published July 23, 2010 Copyright: 2010 Irie et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The work was supported by National Institutes of Health (NIH) grant CA105134, a grant from Sanofi-Aventis, and a gift from Lee Jeans through the Entertainment Industry Foundation to JSB. HYI was supported by 5K08 CA107231 (NIH), the Terri Brodeur Breast Cancer Research Foundation and a Development Pilot Award from the SPORE in Breast Cancer (NIH P50 CA089393). YS was supported by a fellowship from the Department of Defense (DoD) (W81XWH-08-1- 0767), and WH was supported by W81XWH-07-1-0408 (DoD) and R33 CA128625 (NIH/National Cancer Institute [NCI]). GBM and YL were supported by the Kleberg Center for Molecular Markers, P30 CA1667 and P01CA099031. ZW was supported by a grant from the Breast Cancer Research Foundation. JY, KP and LZ were supported by the Specialized Programs of Research Excellence (SPORE) in Breast Cancer (NIH P50 CA089393). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: This work was partially funded by a grant from Sanofi-Aventis to Joan Brugge. We declare that there are no restrictions on sharing any of the data or reagents used in this study in accordance with the policies of PLoS ONE. * E-mail: Joan_Brugge@hms.harvard.edu Introduction Adhesion to extracellular matrix (ECM) provides epithelial cells with critical cues about their environment that are required for their proliferation, survival and tissue organization. Loss of attachment to matrix compromises viability of normal epithelial cells through a variety of mechanisms that help preserve tissue homeostasis and prevent aberrant growth (reviewed in [1], [2]). Detachment from matrix triggers apoptosis, termed anoikis, via both intrinsic and extrinsic death pathways. However, most tumor cells have acquired the ability to resist anoikis and this property is believed to be critical for tumor cell dissemination and survival in altered matrix environments [1], [2]. Genes that have been demonstrated to suppress anoikis also promote metastases in vivo, further supporting a critical role for anoikis regulation in tumorigenesis [3], [4]. Tumor cells adopt several different strategies to evade anoikis including: (1) activation of survival pathways such as those regulated by Erk/MAPK and Akt through oncogenic mutations or constitutive growth factor receptor activation; (2) modulation of expression or activity of anti-apoptotic and pro-apoptotic proteins including Bcl2 family members, and (3) altered expression and engagement of integrins by basement membrane proteins produced via autocrine mechanisms (reviewed in [1], [2]). Large-scale cDNA screens have identified several candidate genes that when overexpressed either alone or in combination PLoS ONE | www.plosone.org 1 July 2010 | Volume 5 | Issue 7 | e11729