In vivo distribution and ex vivo permeation of cyclosporine A prodrug aqueous formulations for ocular application Marta Rodriguez-Aller a , Davy Guillarme a , Mohamed El Sanharawi b , Francine Behar-Cohen b , Jean-Luc Veuthey a , Robert Gurny a, ⁎ a School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, 30, Quai Ernest Ansermet, 1211 Geneva 4, Switzerland b INSERM, UMRS 872 Team 17, Centre de Recherche des Cordeliers, 75006 Paris, France abstract article info Article history: Received 20 December 2012 Accepted 28 April 2013 Available online 3 May 2013 Keywords: Cyclosporine A Water formulation Prodrug In vivo distribution Ex vivo permeation Eye Cyclosporine A is a poorly water-soluble, immunosuppressive drug used to treat a variety of ocular diseases. Its limited solubility makes challenging the development of a cyclosporine A-based eye drop for ocular topical ap- plication. Based on the prodrug strategy, the practically insoluble cyclosporine A was converted into a freely soluble prodrug. Such a water-soluble prodrug made it possible to develop water-based concentrated eye drops. The prodrug formulations were tested for their ex vivo permeation and in vivo distribution at three con- centrations (equivalent to 0.05%, 0.50% and 2.00% w/v cyclosporine A). The ex vivo permeation experiments were performed on corneal and conjunctival epithelia. The in vivo distribution evaluated the total cyclosporine A present in the ocular structures as well as in serum, spleen and cervical lymphatic ganglions. Each prodrug formulation was compared to conventionally used cyclosporine A eye drops at an equivalent concentration. The experimental results showed that the tested eye drops behaved differently. The prodrug formulation was characterized by the following: i) preferential conjunctival penetration, ii) an interesting capacity to create large tissue deposits and iii) a lower risk of systemic complications and immunosuppression. The prodrug aqueous eye drop was demonstrated to be a patient-friendly option for the treatment of ocular diseases requiring high ocular levels of cyclosporine A, pushing the boundaries of the current therapeutic arsenal. © 2013 Elsevier B.V. All rights reserved. 1. Introduction The poor water solubility of active molecules is a major pharmaceuti- cal challenge, as 90% of newly discovered active compounds and 40% of marketed drugs are poorly soluble, according to the biopharmaceutical classification system [1]. Cyclosporine A (CyA) belongs to this group [2]. It is an immunosuppressive peptide that has a variety of applications in ophthalmology, including dry eye syndrome, prevention of corneal graft rejection, blepharitis, rosacea, different types of keratoconjunctivitis or conjunctival graft vs. host disease [3,4]. Nevertheless, the formulation of such a molecule as an eye drop remains challenging. As a consequence, the highest concentration of eye drop conventionally used is 2.00% w/v CyA in oil vehicles [3,4]. When high CyA concentrations are required, oph- thalmologists have no option but systemic administration. Unfortunately, systemic treatments of CyA demand intensive patient monitoring to pre- vent and manage potential systemic side effects, which can be as severe as nephrotoxicity, hepatotoxicity or hypertension [5,6]. The patient's quality of life is drastically changed. The limitations of ocular formulations of CyA are linked to its poor solubilization. CyA requires the use of oily vehicles, which are linked with tolerance issues [7–11] and availability limitations [12–14]. One way to increase the water solubility of CyA was to develop a water- soluble CyA prodrug [2,15–19]. With such a strategy, the “practically in- soluble” CyA molecule was converted into a “freely soluble” CyA prodrug, according to Clarke's classification of drugs [20]. This ester prodrug makes it possible to develop highly concentrated CyA-based ophthalmic formulations within an aqueous vehicle. In addition, this water-based prodrug eye drop avoids tolerance issues linked to the oil vehicles. In the present study, the ex vivo and in vivo permeation and distribu- tion of the newly developed aqueous prodrug formulation were evaluated at different concentrations (0.05% w/v, 0.50% w/v and 2.00% w/v equivalent CyA). Each prodrug formulation was compared to conventional CyA eye drops at the equivalent concentration. In this way, the behaviors of both formulation types could be characterized and compared. Together, ex vivo and in vivo experiments provided es- sential information on CyA and prodrug formulations, allowing a deep understanding of their interactions with the ocular structures and with the rest of the organism. 2. Materials and methods 2.1. Materials The CyA prodrug N-methyl-glucamine salt, was synthesized at the Institute of Chemical Sciences and Engineering of the Ecole Polytechnique Journal of Controlled Release 170 (2013) 153–159 ⁎ Corresponding author. Tel.: +41 223793816. E-mail address: robert.gurny@unige.ch (R. Gurny). 0168-3659/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jconrel.2013.04.019 Contents lists available at SciVerse ScienceDirect Journal of Controlled Release journal homepage: www.elsevier.com/locate/jconrel