Design of a nanostructured lipid carrier intended to improve the treatment of tuberculosis Marina Pinheiro,1,* Ricardo Ribeiro,1,* Alexandre Vieira,1,* Fernanda Andrade,2 Salette Reis1 11UCIBIO, REQUIMTE, Chemistry Department, Faculty of Pharmacy, 2Laboratory of Pharmaceutical Technology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto, Porto, Portugal *These authors contributed equally to this work This work aimed to design, develop, and characterize a lipid nanocarrier system for the selective delivery of rifabutin (RFB) to alveolar macrophages. Lipid nanoparticles, specifically nanostructured lipid carriers (NLC), were synthetized by the high-shear homogenization and ultrasonication techniques. These nanoparticles were designed to exhibit both passive and active targeting strategies to be efficiently internalized by the alveolar macrophages, traffic to the acidified phagosomes and phagolysosomes, and release bactericidal concentrations of the antituberculosis drug intracellularly. NLC that could entrap RFB were prepared, characterized, and further functionalized with mannose. Particles’ diameter, zeta potential, morphology, drug% entrapping efficiency, and drug release kinetics were evaluated. The mannose coating process was confirmed by Fourier transform infrared. Further, the cytotoxicity of the formulations was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay in A549, Calu-3, and Raw 264.7 cells. The diameter of NLC formulations was found to be in the range of 175–213 nm, and drug entrapping efficiency was found to be above 80%. In addition, high storage stability for the formulations was expected since they maintained the initial characteristics for 6 months. Moreover, the drug release was pH-sensitive, with a faster drug release at acidic pH than at neutral pH. These results pose a strong argument that the developed nanocarrier can be explored as a promising carrier for safer and more efficient management of tuberculosis by exploiting the pulmonary route of administration. Keywords: lipid nanoparticles, mannose, nanostructured lipid carriers, rifabutin, tuberculosis, drug delivery systems

Tuberculosis (TB) is still a devastating disease and more people have died of TB than any other infectious diseases throughout the history. The current therapy consists of a multidrug combination in a long-term treatment, being associated with the appearance of several adverse effects. Thus, solid lipid nanoparticles (SLNs) were developed using mannose as a lectin receptor ligand conjugate for macrophage targeting and to increase the therapeutic index of rifampicin (RIF). The developed SLNs were studied in terms of diameter, polydispersity index, zeta potential, encapsulation efficiency (EE) and loading capacity (LC). Morphology, in vitro drug release and differential scanning calorimetry studies, macrophage uptake studies, cell viability and storage stability studies were also performed. The diameter of the SLNs obtained was within the range of 160–250 nm and drug EE was above 75%. The biocompatibility of M-SLNs was verified and the internalization in macrophages was improved with the mannosylation. The overall results suggested that the developed mannosylated formulations are safe and a promising tool for TB therapy targeted for macrophages.