Final Report Summary - NANOGEND (Novel Thermoresponsive Organic Nanogels for Topical Gene Delivery of RNA-Based Drugs) Drug delivery through the human skin is attractive for the clinical treatment of a wide range of acute and chronic skin disorders including psoriasis, skin inflammations and skin cancer. In addition the relatively large and readily accessible skin surface area of 1-2 m2 offers a direct route to the blood system that avoids first pass metabolism and maintains a steady concentration of drug in plasma. It is a non invasive way of drug administration and it avoids painful injections, which in turn generate dangerous medical waste and pose the risk of disease transmission by needle re-use, especially in developing countries. The outer layer of the skin, the stratum corneum (SC), represents a formidable barrier designed to protect the skin, however it seriously impacts the penetration of substances by limiting both topical and transdermal bioavailability. Three basic approaches have been developed to improve bioavailability and increase the range of drugs for which topical and transdermal delivery would be desirable but is currently unfeasible: (i) skin barrier elimination by ablation and abrasion; (ii) stratum corneum perforation by microneedles, electroporation and ultrasounds; (iii) stratum corneum penetration using chemical enhancers or highly adjustable, soft carriers (nanocarriers).The latter option is the least invasive and the most patient complacent, however despite considerable efforts, there is still a lack of suitable vehicles that can efficiently cross the SC. Data in the literature suggest that morphology and physico-chemical characteristics of the materials play a key role and the recent advances in the area of nanomaterials and the emergence of the nanobiotechnology industry have highlighted their significant potential as novel transdermal drug delivery systems using minimally invasive methods. A variety of novel nanomaterials have been developed and studied in the last 15 years, among others liposomes and transferosomes, solid lipid nanoparticles and polymeric nanoparticles, however research has yet to demonstrate enhanced penetration of healthy skin by nanoparticles16,17compared to traditional systems. In addition the lack of analytical tools with high sensitivity, prevents clear tracking of nanoparticles in the systemic route and monitoring of drug release. To be useful in therapeutic applications, nanoparticles must be able to efficiently penetrate the skin barrier, deliver their payload and clear from the body without any adverse side effects. Here we report the preparation of PEG-b-PPS micelles, made of poly(ethylene glycol) as hydrophilic block and poly(ethylene sulfide) as hydrophobic block, with an average diameter size of 15 nm and loaded with flufenamic acid (FFA), and ability to cross the stratum corneum of excised human skin and penetrate the epidermis, without further perturbation of the skin barrier and with consistent delivery of their payload. Due to the low mobility of the hydrophobic block (PPS), which can be considered to be almost frozen when dispersed in an incompatible solvent such as water, the PEG-b-PPS micelles are particularly stable in contrast to classical surfactant micelles and, furthermore, can efficiently encapsulate hydrophobic drugs in their PPS core. The remarkably low critical association concentration (CAC) of PEG-b-PPS micelles, which results in a slow rate of micelles dissociation after dilution, allows retention of loaded drugs for a longer period of time, potentially leading to a higher accumulation of the drug at the target site, a characteristic that is highly valued in therapeutics.The PEG-b-PPS micelles have been designed to bind a 1,8-naphthalimide thio-derivative, N-(pyridin-2-yl-disulfanyl ethyl)-4-(4-methylpiperazin-1-yl)-1,8-naphthalimide (tPNI), as fluorescent sensor covalently attached to the PPS hydrophobic core; the fluorophore displays an ON-OFF switch upon binding with flufenamic acid (FFA), the drug used as a model in this study, allowing monitoring not only of micelles penetration, but also of drug up-loading and release. The results show that the sensitive fluorescent PEG-b-PPS micelles loaded with FFA, penetrate the skin within 24 hours and reach the epidermis, where they deliver an amount of drug that is comparable with the pharmaceutical formulation of FFA .There is currently considerable interest in the skin as an alternative site for drug administration, both for local and systemic treatment and the application of nanotechnology, in particular nanomaterials, to therapeutics may have significant benefit in skin cancer imaging and targeted therapeutics, immunomodulation and vaccine delivery via skin, and antimicrobials and wound healing43,44. The data presented in this study on the use of PEG44PPS22 micelles as transdermal drug delivery system clearly indicate that we are reaching an important stage where these nano-tools can contribute to the clinical translation of nanotechnology to dermatology.