Pulmonary tuberculosis (TB), which is caused by Mycobacterium tuberculosis (Mtb), remains a global pandemic, despite the widespread use of the parenteral live attenuated Bacillus Calmette–Guérin (BCG) vaccine during the past decades. Mucosal administration of next generation TB vaccines has great potential, but developing a safe and efficacious mucosal vaccine is challenging. Hence, understanding the in vivo biodistribution and pharmacokinetics of mucosal vaccines is essential for shaping the desired immune response and for optimal spatiotemporal targeting of the appropriate effector cells in the lungs. A subunit vaccine consisting of the fusion antigen H56 (Ag85B-ESAT-6-Rv2660) and the liposome-based cationic adjuvant formulation (CAF01) confers efficient protection in preclinical animal models. In this study, we devise a novel immunization strategy for the H56/CAF01 vaccine, which comply with the intrapulmonary (i.pulmon.) route of immunization. We also describe a novel dual-isotope (111In/67Ga) radiolabeling approach, which enables simultaneous non-invasive and longitudinal SPECT/CT imaging and quantification of H56 and CAF01 upon parenteral prime and/or i.pulmon. boost immunization. Our results demonstrate that the vaccine is distributed evenly in the lungs, and there are pronounced differences in the pharmacokinetics of H56 and CAF01. We provide convincing evidence that the H56/CAF01 vaccine is not only well-tolerated when administered to the respiratory tract, but it also induces strong lung mucosal and systemic IgA and polyfunctional Th1 and Th17 responses after parenteral prime and i.pulmon. boost immunization. The study furthermore evaluate the application of SPECT/CT imaging for the investigation of vaccine biodistribution after parenteral and i.pulmon. immunization of mice.

Local as well as systemic therapy is often used to treat bacterial lung infections. Delivery of antibiotics to the vascular side of infected lung tissue using lung-targeting microspheres (MS) is a good alternative to conventional administration routes, allowing for localized high levels of antibiotics. This delivery route can also complement inhaled antibiotic therapy, especially in the case of compromised lung function. We prepared and characterized monodisperse poly­(lactic-co-glycolic acid) (PLGA) MS loaded with levofloxacin using a flow-focusing glass microfluidic chip. In vitro characterization showed that the encapsulated LVX displayed a biphasic controlled release during 5 days and preserved its antibacterial activity. The MS degradation was investigated in vitro by cross-sectioning the MS using a focused ion beam scanning electron microscope and in vivo by histological examination of lung tissue from mice intravenously administered with the MS. The MS showed changes in the surface morphology and internal matrix, whereas the degradation in vivo was 3 times faster than that in vitro. No effect on the viability of endothelial and lung epithelial cells or hemolytic activity was observed. To evaluate the pharmacokinetics and biodistribution of the MS, complete quantitative imaging of the 111indium-labeled PLGA MS was performed in vivo with single-photon emission computed tomography imaging over 10 days. The PLGA MS distributed homogeneously in the lung capillaries. Overall, intravenous administration of 12 μm PLGA MS is suitable for passive lung targeting and pulmonary therapy.

The validation of metal–phenolic nanoparticles (MPNs) in preclinical imaging studies represents a growing field of interest due to their versatility in forming predesigned structures with unique properties. Before MPNs can be used in medicine, their pharmacokinetics must be optimized so that accumulation in nontargeted organs is prevented and toxicity is minimized. Here, we report the fabrication of MPNs made of a coordination polymer core that combines In­(III), Cu­(II), and a mixture of the imidazole 1,4-bis­(imidazole-1-ylmethyl)-benzene and the catechol 3,4-dihydroxycinnamic acid ligands. Furthermore, a phenolic-based coating was used as an anchoring platform to attach poly­(ethylene glycol) (PEG). The resulting MPNs, with effective hydrodynamic diameters of around 120 nm, could be further derivatized with surface-embedded molecules, such as folic acid, to facilitate in vivo targeting and multifunctionality. The prepared MPNs were evaluated for in vitro plasma stability, cytotoxicity, and cell internalization and found to be biocompatible under physiological conditions. First, biomedical evaluations were then performed by intrinsically incorporating trace amounts of the radioactive metals 111In or 64Cu during the MPN synthesis directly into their polymeric matrix. The resulting particles, which had identical physicochemical properties to their nonradioactive counterparts, were used to perform in vivo single-photon emission computed tomography (SPECT) and positron emission tomography (PET) in tumor-bearing mice. The ability to incorporate multiple metals and radiometals into MPNs illustrates the diverse range of functional nanoparticles that can be prepared with this approach and broadens the scope of these nanoconstructs as multimodal preclinical imaging agents.