ORIGINAL ARTICLE 9-Aminoacridine-based anticancer drugs target the PI3K/AKT/mTOR, NF-jB and p53 pathways C Guo 1,2 , AV Gasparian 3 , Z Zhuang 4 , DA Bosykh 3 , AA Komar 5 , AV Gudkov 3,6 and KV Gurova 3,6 1 Department of Biochemistry, Case Western Reserve University, Cleveland, OH, USA; 2 Department of Molecular Genetics, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, USA; 3 Cleveland Biolabs Inc., Buffalo, NY, USA; 4 National Institutes of Health, Bethesda, MD, USA; 5 Center for Gene Regulation in Health and Disease, Department of Biology, Cleveland State University, Cleveland, OH, USA and 6 Department of Cell Stress Biology, Roswell Park Cancer Institute, Buffalo, NY, USA Acquisition of a transformed phenotype involves dereg- ulation of several signal transduction pathways contribut- ing to unconstrained cell growth. Understanding the interplay of different cancer-related signaling pathways is important for development of efficacious multitargeted anticancer drugs. The small molecule 9-aminoacridine (9AA) and its derivative, the antimalaria drug quinacrine, have selective toxicity for tumor cells and can simulta- neously suppress nuclear factor-jB (NF-jB) and activate p53 signaling. To investigate the mechanism underlying these drug activities, we used a combination of two- dimensional protein separation by gel electrophoresis and mass spectrometry to identify proteins whose expression is altered in tumor cells by 9AA treatment. We found that 9AA treatment results in selective downregulation of a specific catalytic subunit of the phosphoinositide 3-kinase (PI3K) family, p110c. Further exploration of this observation demonstrated that the mechanism of action of 9AA involves inhibition of the prosurvival AKT/ mammalian target of rapamycin (mTOR) pathway that lies downstream of PI3K. p110c translation appears to be regulated by mTOR and feeds back to further modulate mTOR and AKT, thereby impacting the p53 and NF-jB pathways as well. These results reveal functional interplay among the PI3K/AKT/mTOR, p53 and NF-jB pathways that are frequently deregulated in cancer and suggest that their simultaneous targeting by a single small molecule such as 9AA could result in efficacious and selective killing of transformed cells. Oncogene (2009) 28, 1151–1161; doi:10.1038/onc.2008.460; published online 12 January 2009 Keywords: proteomics; quinacrine; cell death; PI3 kinase-g; PTEN Introduction Tumor cells frequently acquire prosurvival mechanisms leading to deregulation of major stress response path- ways. Suppression of p53 and constitutive activation of nuclear factor-kB (NF-kB) are among the most com- mon properties of tumor cells and are, therefore, considered potential targets for therapeutic intervention aimed at selective elimination of such cells. In addition, cell transformation often involves activation of prosur- vival pathways contributing to unconstrained cell growth under normally restrictive conditions, including AKT/mammalian target of rapamycin (mTOR) and phosphoinositide 3-kinase (PI3K) pathways. There are numerous indications of interaction between these pathways, which suggests that their simultaneous pharmacological modulation would provide enhanced anticancer efficacy (Mayo and Donner, 2001; Sizemore et al., 2002; Levine et al., 2006; Budanov and Karin, 2008). p53 is a key tumor suppressor that plays an undisputed role in cancer prevention. p53 tumor suppressive activity stems from its ability to respond to a variety of stresses and trigger cell cycle arrest, apoptosis or senescence, thereby protecting against malignant transformation (Bensaad and Vousden, 2005). Understanding the mechanisms responsible for functional inactivation of wild-type p53 in human tumors (for example, overexpression of natural antago- nists of p53 such as Mdm2 or the viral E6 protein) helps to define prospective targets for treating cancer by restoring p53 function (Gudkov, 2005). Like p53, NF-kB is a major sensor of cell stress. Activation of the NF-kB signaling pathway results in inhibition of apoptosis by induction of antiapoptotic genes and/or suppression of proapoptotic genes (Kucharczak et al., 2003). Consistent with its antiapoptotic activity, NF-kB has been reported to be constitutively active in numerous tumors, reducing their sensitivity to proapoptotic stresses, including those associated with cancer treatment (Karin and Gretin, 2005). p53 and NF- kB antagonize each other and the balance in activity of the two pathways determines whether a cell proliferates or undergoes apoptosis in stress conditions. Received 24 October 2008; accepted 20 November 2008; published online 12 January 2009 Correspondence: Dr AV Gudkov and Dr KV Gurova, Department of Cell Stress Biology, BLSC L3-301, Roswell Park Cancer Institute, Elm and Carlton Streets, Buffalo, NY 14263, USA. E-mails: andrei.gudkov@roswellpark.org and katerina.gurova@roswellpark.org Oncogene (2009) 28, 1151–1161 & 2009 Macmillan Publishers Limited All rights reserved 0950-9232/09 $32.00 www.nature.com/onc