Current treatment options for bone-related disorders rely mostly on the systemic administration of therapeutic agents with low solubility/intracellular bioavailability and high pharmacokinetic variability, leading to major off-target side effects. Subsequently, there is an unmet balance between drug delivery and its clinical efficacy. To overcome these issues, nanoparticle-delivery systems might be promising carriers of hydrophobic agents. Therefore, we propose to design a nanoparticle-delivery system for the intracellular controlled-release of small hydrophobic drugs for bone-disorders therapies. To test our system, we selected Dexamethasone as a hydrophobic/lipophilic drug model, known to play a dual regulatory role in bone metabolism/formation depending on dosage and treatment length. Accordingly, a higher (10−6 M) or lower (10−7 M) concentration of Dexamethasone was loaded into two distinct polymeric-nanoparticles, mPEG-PTMC-copolymer and a FDA-approved-polymer PLGA. The bio-functionality of the developed nanoparticles was further analysed in vitro, using MC3T3-E1 pre-osteoblasts and primary bone marrow stromal cells (isolated from wild-type C25BL/6 mice) in contact with Dexamethasone-loaded nanoparticles. Our studies demonstrated that nanoparticles were able to rapidly internalise within 4 hours-of-incubation, modulating the response of osteoblasts to Dexamethasone accordingly with dosage delivered. In fact, the release of 10−6 M Dexamethasone by loaded-nanoparticles inhibited significantly osteoblast metabolic activity when compared to non-treated cells (P<0.001), while the release of 10−7 M Dexamethasone promoted a 2-fold increase in osteoblast activity (P<0.001), which was sustained for 14 days. Moreover, the release of 10−7 M Dexamethasone also induced osteoblast differentiation, as seen by a trend towards increased alkaline phosphatase activity (P=0.07) at day 7-of-incubation and increased osteocalcin levels (P<0.001) as well as calcium deposition (matrix mineralization; P<0.001) at day 14-of-incubation. In conclusion, we describe for the first time a promising therapeutic strategy for the intracellular delivery of small hydrophobic agents based on polymeric-nanoparticles, demonstrating its in vivo therapeutic potential for the delivery of drugs targeting downstream-signalling pathways involved in bone-related disorders.
Disclosure: The authors declared no competing interests. This work was financed by FEDER funds through the Programa Operacional Factores de Competitividade COMPETE and by Portuguese funds through the Fundação para a Ciência e a Tecnologia FCT in the framework of the financed projects PEst-C/SAU/LA0002/2013 and PTDC/BIM-MED/1047/2012.