Published: April 29, 2011 r2011 American Chemical Society 1249 dx.doi.org/10.1021/ci100501d | J. Chem. Inf. Model. 2011, 51, 1249–1258 ARTICLE pubs.acs.org/jcim Rational Design of New Class of BH3-Mimetics As Inhibitors of the Bcl-x L Protein Marta Pinto, †,‡ Maria del Mar Orzaez, § Laura Delgado-Soler, † Juan J. Perez, ‡ and Jaime Rubio-Martinez* ,† † Department of Physical Chemistry, University of Barcelona (UB), and Instituto de Química Te orica y Computacional (IQTCUB), Barcelona, Spain ‡ Department of Chemical Engineering, Polytechnic University of Catalonia (UPC), ETSEIB, Catalonia, Spain § Department of Medicinal Chemistry, Centro de Investigaci on Príncipe Felipe, and Instituto de Biomedicina de Valencia, CSIC, Valencia, Spain b S Supporting Information ’ INTRODUCTION Apoptosis or programmed cell death is essential during embryogenesis and cellular homeostasis and plays an important role in the defense against pathogens. Defects in apoptotic pathways contribute to a number of human diseases, such as cancer and autoimmune or neurodegenerative diseases. 1,2 Apop- tosis is orchestrated by a group of intracellular cysteinyl aspartate proteases called caspases, 3 synthesized as inactive zymogens that are activated through two major interconnected pathways: 4,5 an extrinsic pathway, activated by ligation of the so-called cell death receptors, and an intrinsic pathway, caused by perturbation of the mitochondria and regulated by the Bcl-2 family of proteins, which is the focus of the present report. To date, at least 20 Bcl-2 family members have been identified in mammalian cells, including proteins that promote apoptosis (hereafter referred as pro-apoptotic) and those that prevent it (hereafter referred as anti-apoptotic). All these proteins are characterized by exhibiting at least one of the four highly conserved so-called Bcl-2 homology domains (hereafter referred to as BH1-BH4), that are useful to classify these proteins into three groups: (1) proteins, such as Bcl-2, Bcl-x L , Mcl-1, and Bfl-1, all of which exhibit anticell death and conserve the four highly conserved domains BH1-BH4; (2) proapoptotic multidomain members, such as Bax, Bak, or Bok, which share sequence homology in the BH1, BH2, and BH3 domains, and finally, (3) the BH3-only proteins such as Bid, Bim, Bmf, Bik, Noxa, Puma, Bad, and Hrk, which contain only a BH3 domain essential for their killing function. 6À10 The relative amount between pro- and anti-apoptotic proteins determines the susceptibility of the cell to apoptosis. Indeed, in many tumor cells Bcl-x L is overexpressed, impairing cell cap- ability to undergo apoptosis and showing resistance to conven- tional anticancer therapies. Therefore, inhibition of this protein or reduction of its level modulates the drug resistance of tumor cells by increasing their susceptibility to undergo apoptosis. The anticell death activity of the anti-apoptotic proteins is due, at least in part, to their dimerization with proapoptotic members of the Bcl-2 family. Structural studies reveal that, in response to pro- survival stimuli, anti-apoptotic proteins undergo a conformational change that leads to the formation of a hydrophobic groove on their surface where the amphipathic R-helical BH3 domain of the pro- apoptotic protein binds. 11À14 By this process overexpressed pro- survival proteins sequester pro-apoptotic ones compromising cell capability to undergo apoptosis. Interestingly, the BH3 domain fragments of pro-apoptotic proteins exhibit high affinity for the pro- survival proteins and play the role of antagonist of these proteins. Indeed, binding of these peptides to the pro-survival proteins has Received: December 21, 2010 ABSTRACT: The Bcl-2 family of proteins plays an important role in the intrinsic pathway of cell apoptosis. Overexpression of pro- survival members of this family of proteins is often associated with the development of many types of cancer and confers resistance against conventional therapeutic treatments. Accordingly, antag- onism of its protective function has emerged as an encouraging anticancer strategy. In the present work, we use a pharmacophore for describing interaction between the BH3 domain of different pro-apoptotic members and the pro-survival protein Bcl-x L in order to identify new lead compounds. In the strategy followed in the present work, the pharmacophore was derived from molecular dynamics studies of different Bcl-x L /BH3 complexes. This pharmacophore was later used as query for 3D database screening. Hits obtained from the search were computationally assessed, and a subset proposed for in vitro testing. Two of the 15 compounds assayed were found able to disrupt the Bcl-x L /Bak(BH3) complex with IC 50 values in the lower micromolar range. Finally, docking studies were performed to explore the binding mode of these compounds to Bcl-x L for further modifications.