Cord Blood Stem Cells Inhibit Epidermal Growth Factor Receptor Translocation to Mitochondria in Glioblastoma Venkata Ramesh Dasari1, Kiran Kumar Velpula1, Kiranmai Alapati1, Meena Gujrati3, Andrew J. Tsung2,4* 1 Departments of Cancer Biology and Pharmacology, University of Illinois College of Medicine at Peoria, Peoria, Illinois, United States of America, 2 Department of Neurosurgery, University of Illinois College of Medicine at Peoria, Peoria, Illinois, United States of America, 3 Department of Pathology, University of Illinois College of Medicine at Peoria, Peoria, Illinois, United States of America, 4 Illinois Neurological Institute, University of Illinois College of Medicine at Peoria, Peoria, Illinois, United States of America Abstract Background: Overexpression of EGFR is one of the most frequently diagnosed genetic aberrations of glioblastoma multiforme (GBM). EGFR signaling is involved in diverse cellular functions and is dependent on the type of preferred receptor complexes. EGFR translocation to mitochondria has been reported recently in different cancer types. However, mechanistic aspects of EGFR translocation to mitochondria in GBM have not been evaluated to date. Methodology/Principle Findings: In the present study, we analyzed the expression of EGFR in GBM-patient derived specimens using immunohistochemistry, reverse-transcription based PCR and Western blotting techniques. In clinical samples, EGFR co-localizes with FAK in mitochondria. We evaluated this previous observation in standard glioma cell lines and in vivo mice xenografts. We further analyzed the effect of human umbilical cord blood stem cells (hUCBSC) on the inhibition of EGFR expression and EGFR signaling in glioma cells and xenografts. Treatment with hUCBSC inhibited the expression of EGFR and its co-localization with FAK in glioma cells. Also, hUCBSC inhibited the co-localization of activated forms of EGFR, FAK and c-Src in mitochondria of glioma cells and xenografts. In addition, hUCBSC also inhibited EGFR signaling proteins in glioma cells both in vitro and in vivo. Conclusions/Significance: We have shown that hUCBSC treatments inhibit phosphorylation of EGFR, FAK and c-Src forms. Our findings associate EGFR expression and its localization to mitochondria with specific biological functions in GBM cells and provide relevant preclinical information that can be used for the development of effective hUCBSC-based therapies. Citation: Dasari VR, Velpula KK, Alapati K, Gujrati M, Tsung AJ (2012) Cord Blood Stem Cells Inhibit Epidermal Growth Factor Receptor Translocation to Mitochondria in Glioblastoma. PLoS ONE 7(2): e31884. doi:10.1371/journal.pone.0031884 Editor: Antonio Paolo Beltrami, University of Udine, Italy Received August 5, 2011; Accepted January 20, 2012; Published February 14, 2012 Copyright: 2012 Dasari 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: This research is supported by a grant from Illinois Neurological Institute (to AJT). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: Andrew.J.Tsung@INI.org Introduction Epidermal growth factor receptor (EGFR) expression in glioblastoma multiforme (GBM) has both diagnostic and prog- nostic significance [1–4]. EGFR signaling plays an important role in the formation and tumorigenicity of malignant gliomas, and EGFR overexpression can be found in 40 to 50 percent of patients with GBM [5]. Multiple histopathological and genetic studies, together with recent large-scale cancer gene sequencing efforts, have identified EGFR and its downstream signaling networks as commonly deregulated components in the primary GBM tumors [1,6,7]. Furthermore, substantial experimental evidence support a causal role for aberrant EGFR signaling in cancer pathogenesis and resistance to treatment [8]. GBM show very high resistance to treatment with radiation and chemotherapy and aberrant EGFR signaling contributes to this resistance. Thus, although initial attempts to target the EGFR have not been effective in GBM [9], EGFR remains an attractive target for therapeutic intervention in GBM. Binding of the ligand epidermal growth factor (EGF) to EGFR triggers receptor dimerization and activates tyrosine kinase, which results in phosphorylation of the tyrosine residues of the receptor and ultimately initiates an array of signaling events [10,11]. Besides the generation of multiple signal transduction events, the activation of EGFR by ligand also dramatically changes the cellular localization of receptors by accelerating internalization of EGFR through clathrin-dependent and -independent endocytic pathways. EGFR has been found in caveoli, Golgi, endoplasmic reticulum, lysosome-like structures, nuclear envelopes, nuclei and within mitochondria [12–14]. EGFR translocates to mitochondria through interaction with proteins such as cytochrome c oxidase (Cox) subunit II [12]. Demory et al. [15] hypothesized that EGF stimulation would lead to EGFR and c-Src activation followed by translocation of these molecules to the mitochondria, phosphor- ylation of CoxII by EGFR and/or c-Src, reduction in oxidative ATP and free radical production, and an increase in cell viability. However, mechanistic aspects of EGFR translocation to mito- chondria in GBM have not been evaluated to date. EGFR has been strongly implicated in the biology of human epithelial malignancies, with therapeutic applications in cancers of the colon, head and neck, lung, and pancreas. Accordingly, the targeting of EGFR has been intensely pursued and has resulted in the PLoS ONE | www.plosone.org 1 February 2012 | Volume 7 | Issue 2 | e31884