Dual System for the Central Nervous System Targeting and Blood-Brain Barrier Transport of a Selective Prolyl Oligopeptidase Inhibitor Meritxell Teixid o, 1 Esther Zurita, 1 Laura Mendieta, 1 Benjam  ı Oller-Salvia, 1 Roger Prades, 1 Teresa Tarrag o, 1 Ernest Giralt 1,2 1 Institute for Research in Biomedicine (IRB Barcelona), Barcelona Science Park, Baldiri Reixac 10, Barcelona 08028, Spain 2 Department of Organic Chemistry, University of Barcelona, Mart  ı i Franque `s 1-11, Barcelona 08028, Spain Received 4 February 2013; accepted 19 April 2013 Published online 26 July 2013 in Wiley Online Library (wileyonlinelibrary.com). DOI 10.1002/bip.22275 ABSTRACT: Less than 2% of all potential neurotherapeutics cross the blood-brain barrier (BBB). Here, we sought to build a construct with the capacity to cross this barrier, to behave as a chemical delivery system, and, once inside the central nervous system, to be transformed and then act as an enzyme inhibitor. With all this in mind, here, we describe the entire process to obtain such a compound, from the initial candidate selection to preparation of the com- pound library and posterior evaluation and final selection of the most promising candidates in terms of selectivity, serum stability, and BBB-transport. V C 2013 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 100: 662–674, 2013. Keywords: blood-brain barrier; drug delivery; central nervous system; diketopiperazine; enzyme inhibitor This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by email- ing the Biopolymers editorial office at biopolymers@wiley.com INTRODUCTION S everal severe health disorders require treatment of the brain. These include neurodegenerative diseases, such as Parkinson’s and Alzheimer’s, but also other central nervous system (CNS) diseases, such as schiz- ophrenia, epilepsy, and bipolar disorder. Cancer, human immunodeficiency virus (HIV), and even certain aspects of obesity are also considered cerebral pharmaceuti- cal targets. During recent years, we have devoted intensive research effort to the field of drug delivery to the brain. 1–5 The delivery of pharmacological agents to the brain is not an easy task. In fact, the scientific community has long dreamt about this challenge. In 1966, a film called “The Fantastic Voy- age” addressed the issue of sending a shuttle to treat the brain. This was a complex journey as the drugs directed toward tar- gets inside the brain had to overcome the blood-brain barrier (BBB). The discovery of the BBB in the late 19th century stemmed from Paul Ehrlich’s 6 observation that intravenously administered dyes stain all organs except the brain. In 1913, Edwin Goldman 7 hypothesized that cerebral capillaries provide the anatomical basis for this physiological barrier. In the late 1960s, this hypothesis was confirmed by electron micros- copy. 8,9 The BBB is present in all vertebrate brains and acts as a natural defense to protect this organ. In fact, this barrier is re- sponsible for more than 98% of all potential neurotherapeutics failing to reach the brain. 10 While blood vessels have pores between cells, brain capillaries have tight junctions, thereby avoiding the paracellular pathway. Thus, all transport occurs through endothelial cells. Various transport mechanisms that allow passage across these cells ensure that certain nutrients reach the brain. Given the impermeability of the BBB, these transport mechanisms may prove effective in delivering drugs to the brain. One of the BBB-shuttles that we have been working with in recent years is based on a diketopiperazine (DKP) scaf- fold, modified to attach cargos and carry them across the Additional Supporting Information may be found in the online version of this article. Correspondence to: Meritxell Teixid o; e-mail: meritxell.teixido@irbbarcelona.org or Ernest Giralt; e-mail: ernest.giralt@irbbarcelona.org Contract grant sponsor: MCI-FEDER Contract grant number: Bio2008-00799 Contract grant sponsor: Generalitat de Catalunya Contract grant numbers: XRB and 2009SGR-1005 Contract grant sponsor: MARAT  O TV3 V C 2013 Wiley Periodicals, Inc. 662 PeptideScience Volume 100 / Number 6