Diagnostics and vaccines for Visceral Leishmaniasis

This project is a pump-priming award to establish collaborations between UK and Brazilian researchers. Introduction: Leishmaniasis is a parasite disease that is endemic in 98 countries worldwide, with the burden of infections in poor and disadvantaged communities in Brazil and India. It is the second most important parasitic infection globally after malaria, with approximately 700,000 – 1 million cases reported. The most serious presentation is visceral leishmaniasis, which has a mortality rate approaching 100% for untreated patients. The key bottlenecks in controlling leishmaniasis globally are 1) rapid identification of infected patients at the point-of-care (POC), 2) the urgent need for vaccines for human infection and better canine vaccines and 3) better treatments to replace those currently used that have high levels of toxicity and side effects. In the current collaboration we propose to try and establish and examine some key aspects of leishmania infction and control -: 1) Diagnosis: improving current recombinant antigen to capture circulating antibodies. a) Experimental determination of antigen-antibody binding affinities. At FIOCRUZ, using bioassays and drug screening facility run by Prof. Floriano Silva, we will determine the experimental binding affinities by microscale thermoforesis and isothermal titration calorimetry of antigen-antibody pairs provided by Dr. Christodoulides. b) Molecular modelling with alternative antigens. Next, at FIOCRUZ, we will use computer-assisted molecular modelling of the protein antigens and antibodies to better understand the structural basis for the antigen-antibody interactions and employ that information to design improved antigens to enhance assay sensitivity. 2 & 3) Better treatments and new vaccines: our hypothesis for developing new treatments is based on the fact that production of nitric oxide (NO) is important for protection against leishmaniasis either through stimulation by vaccination or delivery via antiparasitic drugs. Similar to NO, silver nanoparticles (AgNPs) present a broad spectrum of applications as antibacterial, antifungal, and antiviral agent. Recently, Dr Seabra´s group first demonstrated that the combination of NO donors and AgNPs has a potent anticancer and antimicrobial effects. In this proposal we aim to combine AgNPs with NO by chemical functionalization of the surface of the nanoparticles with NO donors to have synergistic effects against Leishmania. The recent advances in nanotechnology have brought several advantages of nanoparticles as a nanocarrier-based drug delivery system to overcome conventional treatments of leishmaniasis. The drug-loaded nanoparticles promote a gradual and sustained drug release improving biodistribution, stability, solubility and decreasing undesirable cytotoxicity. Within the context of this small grant we plan to join the expertise of the UK PI on Leishmania with Dr. Seabra’s know-how on AgNPs and Dr. Silva-Jr’s experience on cellular and molecular assays to pursue the following goals: c) Measure internalization of silver nanoparticles by the protozoa at UFABC by the Single Particle ICP-MS technique. d) Determine nitrosylated proteins through western blot (commercial antibodies) and high content analysis to locate intracellularly and temporally, using a novel technique at FIOCRUZ (high content analysis (HCA), which images biological samples under a fluorescence, confocal or widefield microscope followed by computational processing of the images to extract features that can be correlated to biologically relevant phenotypes). The large image datasets combined with the power of statistical analysis make HCA an important methodology to unveil new biological knowledge.