Published: June 27, 2011 r2011 American Chemical Society 5882 dx.doi.org/10.1021/jo2007785 | J. Org. Chem. 2011, 76, 5882–5894 FEATURED ARTICLE pubs.acs.org/joc β-Cyclodextrin-Based Polycationic Amphiphilic “Click” Clusters: Effect of Structural Modifications in Their DNA Complexing and Delivery Properties Alejandro M endez-Ardoy, † Nicolas Guilloteau, ‡ Christophe Di Giorgio, ‡ Pierre Vierling,* ,‡ Francisco Santoyo-Gonz  alez, § Carmen Ortiz Mellet,* ,† and Jos  e M. García Fern  andez* ,|| † Departamento de Química Org  anica, Facultad de Química, Universidad de Sevilla, Profesor García Gonz  alez 1, E-41012 Sevilla, Spain ‡ LCMBA UMR 6001CNRS - Universit  e de Nice Sophia Antipolis 28, Avenue de Valrose, F-06108 Nice, France § Departamento de Química Org  anica, Facultad de Ciencias, Instituto de Biotecnología, Universidad de Granada, E-18071 Granada, Spain ) Instituto de Investigaciones Químicas, CSIC - Universidad de Sevilla, A2m erico Vespucio 49, Isla de la Cartuja, E-41092 Sevilla, Spain b S Supporting Information ’ INTRODUCTION Gene therapy is a promising strategy for the treatment of a broad range of ailments including various types of cancer and cardiovascular, monogenic, and infectious diseases. The success of this approach is strongly dependent on the development of safe and efficient gene delivery systems that can provide protection of the therapeutic gene and effectively deliver exogenous genetic material into specific cell types. Much attention has been focused initially on viral-based delivery systems. However, their limited DNA carrying capacity, expensive cost, and safety concerns such as immunogenic response, toxicity or oncogenicity are major limitations inherent to these vectors. 1 During the last two decades, nonviral gene delivery systems have emerged as an attractive alternative. 2 A large diversity of (poly)cationic lipids, polymers, and dendrimers has been explored for gene transfer applications, and their use has moved from in vitro transfection to clinical gene therapy trials. Unfortunately, low efficiency and poor selectivity compared to their viral counterparts severely limit their applica- tion range. 3 Understanding the mechanisms involved in cell and systemic traffic of vector:plasmid DNA (pDNA) complexes and elucida- tion of the relationships between vector structure and transfec- tion efficiency is essential for the rational design of efficient nonviral gene delivery systems. Facial amphiphiles consisting of a nanometric platform such as fullerenes, calix[4]resorcinarenes (resorcarenes), calixarenes, or cyclodextrins (CD), onto which a variety of cationic groups and lipophilic moieties can be installed with a precise spatial orientation, have been shown to be particularly well-suited for this purpose. 4 As for classical cationic amphiphiles, facial cationic amphiphiles are able to form orga- nized supramolecular structures with DNA. The resulting com- plexes can display further fusogenic, 5 permeation, 6 molecular recognition, 7 and/or transfection properties. 8 Most interestingly, homogeneity can be preserved at the molecular level in structu- rally related series of compounds by implementing selective Received: April 17, 2011 ABSTRACT: Monodisperse facial amphiphiles consisting of a β-cyclo- dextrin (βCD) platform exposing a multivalent display of cationic groups at the primary rim and bearing hydrophobic chains at the secondary oxygens have been prepared by implementing two very robust “click” methodologies, namely cuprous cation-catalyzed azideÀalkyne cycload- dition (CuAAC) and thiourea-forming reaction. Most interestingly, the use of solid-supported Cu(I) catalysts was found to be very well suited for multiple CuAAC while facilitating purification of the C 7 - symmetric macromolecular triazole adducts. The strategy is compatible with molecular diversity-oriented approaches, which has been exploited to generate a small library of click polycationic amphiphilic CDs (paCDs) for assessing the influence of structural modifications in the ability to complex, compact, and protect pDNA and the efficiency of the resulting paCD:pDNA nanocomplexes (CDplexes) to deliver DNA into cells and promote transfection. The results indicate that fine-tuning the hydrophilic/hydrophobic balance is critical to achieve optimal self-assembling properties and stability of the resulting CDplexes in saline- and serum-containing media. Triazole-type paCDs were, in general, less efficient in promoting gene transfection than thiourea-type derivatives. Nevertheless, the current body of results support that the “dual click” approach implying sequential CuAAC and thiourea-forming reactions represents a versatile strategy to optimize the gene delivery capabilities of cyclodextrin- based facial amphiphiles.