Inorganic theranostic nanoparticles for Tuberculous Meningitis

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Tuberculous meningitis (TBM) is the most severe form of tuberculosis infection with very high mortality and accountsfor 2-5% of all tuberculosis (TB) cases among children and HIV-positive adults, causing permanent neurologicalconsequences and disability. There is an urgent clinical need to develop vehicles to deliver antimicrobials/antiinflammatoriesdirectly inside the brain to treat and diagnose early stage TBM.The PhD research project will focus on 4 tasks: (1) synthesis of theranostics nanoparticles (NPs) with controlled size(<100 nm) for optimal BBB crossing. (2) Decoration of NPs with pH responsive polymer which allows NPs to be absorbedand pass into the blood stream. (3) NPs physicochemical characterisation. (4) In vitro testing of NPs transcytosisformulation.Task 1: Synthesis of NPs. The PhD student will design and develop spherical (MSNs) and flower (MSNFs) shapemesoporous silica NPs with controlled size (<100 nm) containing first line antibiotics for optimal BBB crossing. The silicanetwork will be obtained by a modified Stöber method. Different concentrations of therapeutic ions (Ce, and Fe - forMRI imaging) will be incorporated into MSNs using a post grafting method.Task 2: Decoration of NPs with pH responsive polymer. A double polymeric shell will be synthesised in order toenable the oral administration of the new therapeutic NPs. MSNs/MSNFs will be decorated with pH responsive polymer(Eudragit E100) which is soluble and swells at gastric pH (up to 5), allowing NPs to be absorbed and pass into the bloodstream. Moreover, a permeable pH independent polymer (Eudragit RL 100) will be used to decorate MSNs and MSNFs.Eudragit RL 100 avoids drug release in the stomach and in the gut. COOH-MSNs and COOH-MSNFs will befunctionalised, using carbodiimide as coupling reagents, with the L-Dopa molecule that is shown to strongly improvetheir BBB crossing.Task 3: MSNs and MSNFs physicochemical characterisation. Ion release kinetics of the MSNs/MSNFs@CeO2-Fe3O4 in different biological media will be performed by ICP-MS. The NPs will be characterised using a range oftechniques including TEM-EDX, NMR, DLS, SAXS/WAXS and BET and HPLC. The magnetic properties of the NPs willbe studied by using a SQUID magnetometer. XPS and EELS will be used to measure the oxidation state of the ceria indifferent media.Task 4: In vitro testing of MSNs/MSNFs@CeO2-Fe3O4 transcytosis formulations.The most promising MSNs/MSNFs@CeO2-Fe3O4 systems will be selected for testing in vitro. Cell lines and primaryastrocytes, microglia, and neurons will be cultured with selected MSNs/MSNFs@CeO2-Fe3O4. The toxicity of the newsystem to healthy neuronal (astrocyte, microglia, neurons) and infected THP-1 microglial cells will be tested by usingMTT and LDH assays. ROS production will also be measured and correlated to the oxidation state of the nanoceria.Inflammation will be assayed by measuring cytokine release. The antibacterial properties of the NPs system will bemeasured using a colony forming assay.To investigate the ability of the MSNs/MSNFs@CeO2-Fe3O4 to cross the BBB and, if necessary, adjust the nano-drugformulation, a convergence of multi-disciplinary in vitro studies, using monocultures and specialised, unique co-culturemodels, of TBM, microglia and brain endothelial cell monolayers being developed at the Crick by Prof. Robert Wilkinsontranswell model of the TBM infected BBB will be used. In both the barrier and diseased cells, a range of cell andmolecular biological processes will be monitored, including nanocarrier drug efficacy, bioreactivity, particle uptake,transformation, particle degradation/products of degradation, translocation, and function and BBBpermeability/disruption.