Cationic Supramolecular Vesicular Aggregates for Pulmonary Tissue Selective Delivery in Anticancer Therapy Mariano Licciardi, [a] Donatella Paolino, [b, c] Nicolò Mauro, [a] Donato Cosco, [c, d] Gaetano Giammona, [a, e] Massimo Fresta, [c, d] Gennara Cavallaro, [a] and Christian Celia* [f, g] Introduction In recent last years, innovative biopolymers and colloidal nano- particles, such as liposomes, were self-assembled to synthesize a new generation of supramolecular vesicular aggregates (SVAs), which can be used in pharmaceutics [1, 2] and nanomedi- cines. [3, 4] Liposome/polymer-based SVAs provide several advan- tages over conventional vesicular and polymeric nanoparticles. In particular, these innovative nanoconstructs show the main advantages of biocompatible polymers, for example, linear and/or branched polymers with several appended groups that can be easily activated through a quick reaction to conjugate several targeted molecules, [5] and liposomes, which are safe, biodegradable, biocompatible, and capable of co-delivering drugs with different physicochemical properties. [6–8] Recently, we synthesized innovative SVAs made from a,b- poly(aspartyl hydrazide) (PAHy) to deliver chemotherapeutics and investigated their biopharmaceutical features to achieve suitable nanoformulations for potential clinical treatment. [1–3] In particular, PAHy copolymer, a water-soluble, biocompatible, nontoxic, and non-antigenic polymer (previously used to syn- thesize therapeutic micelles, [9] innovative scaffolds for tissue engineering, [10] polycationic complexes for gene delivery [11] and zwitterionic polypeptide nanogels [12] ) was self-assembled in pre-formulated liposomes by its hydrophobic butyric moieties (PAHy-C 4 ) to form SVAs. The butyric residue anchored copoly- mers to the phospholipid bilayer and exposed the hydrophilic chains toward the external aqueous environment. [13] PAHy-C 4 copolymers covering the surface of SVAs show physicochemical properties similar to polyethylene glycol (PEG) derivatives conjugated to phospholipids, which are used to synthesize stealth liposomes. PAHy-C 4 copolymers can modify the surface charge of SVAs [14] as well as in vivo targeting and biodistribution. [1] In fact, (carboxypropyl)trimethylammonium hydrochloride (CPTA) moieties, which are conjugated to the backbone structure of PAHy-C 4 and form PAHy-C 4 –CPTA, are used to synthesize cationic SVAs. The resulting cationic nano- carriers can strongly interact with the surface of various tumor cells. [14] The conjugation of folate to create PAHy-C 4 –folate co- polymers forms SVAs–folate, which selectively target cancer cells overexpressing folate receptors. [2] Surface modifications of The biopharmaceutical properties of supramolecular vesicular aggregates (SVAs) were characterized with regard to their physicochemical features and compared with cationic lipo- somes (CLs). Neutral and cationic SVAs were synthesized using two different copolymers of poly(aspartyl hydrazide) by thin- layer evaporation and extrusion techniques. Both copolymers were self-assembled in pre-formulated liposomes and formed neutral and cationic SVAs. Gemcitabine hydrochloride (GEM) was used as an anticancer drug and loaded by a pH gradient remote loading procedure, which significantly increased drug loading inside the SVAs. The resulting average size of the SVAs was 100 nm. The anticancer activity of GEM-loaded neutral and cationic SVAs was tested in human alveolar basal epithelial (A549) and colorectal cancer (CaCo-2) cells. GEM-loaded cation- ic SVAs increased the anticancer activity in A549 and CaCo-2 cells relative to free drug, neutral SVAs, and CLs. In vivo biodis- tribution in Wistar rats showed that cationic SVAs accumulate at higher concentrations in lung tissue than neutral SVAs and CLs. Cationic SVAs may therefore serve as an innovative future therapy for pulmonary carcinoma. [a] Prof. M. Licciardi, Prof. N. Mauro, Prof. G. Giammona, Prof. G. Cavallaro Laboratory of Biocompatible Polymers, Biological, Chemical and Pharma- ceutical Sciences and Technologies Department (STEBICEF), University of Palermo, Via Archirafi 32, 90123 Palermo (Italy) [b] Prof. D. Paolino Department of Experimental and Clinical Medicine, Building of BioSciences, University of Catanzaro “Magna Græcia”, V.le Europa s.n.c., 88100 Germaneto (Italy) [c] Prof. D. Paolino, Dr. D. Cosco, Prof. M. Fresta Interregional Research Center for Food Safety & Health (IRCFSH), Building of BioSciences, University of Catanzaro “Magna Græcia”, V.le Europa s.n.c., 88100 Germaneto (Italy) [d] Dr. D. Cosco, Prof. M. Fresta Department of Health Sciences, University of Catanzaro “Magna Græcia”, Building of BioSciences, V.le Europa s.n.c., 88100 Germaneto (Italy) [e] Prof. G. Giammona Mediterranean Center for Human Advanced Biotechnologies (Med-Chab), Viale delle Scienze Ed. 18, 90128 Palermo (Italy) [f] Dr. C. Celia Department of Pharmacy, University of Chieti – Pescara “G. d’Annunzio”, Via dei Vestini 31, 66100 Chieti (Italy) E-mail : c.celia@unich.it [g] Dr. C. Celia Department of Nanomedicine, Houston Methodist Research Institute, 6670 Bertner Avenue, Houston, TX 77030 (USA) Supporting information and the ORCID identification number(s) for the author(s) of this article can be found under http://dx.doi.org/10.1002/ cmdc.201600070. ChemMedChem 2016, 11, 1 – 12  2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim 1 & These are not the final page numbers! ÞÞ These are not the final page numbers! ÞÞ Full Papers DOI: 10.1002/cmdc.201600070