Towards Translational ImmunoPET/MR Imaging of Invasive Pulmonary Aspergillosis: The Humanised Monoclonal Antibody JF5 Detects Aspergillus Lung Infections In Vivo

This is the final published versionAvailable from Ivyspring International Publisher via the DOI in this recordInvasive pulmonary aspergillosis (IPA) is a life-threatening lung disease of hematological malignancy and bone marrow transplant patients caused by the ubiquitous environmental fungus Aspergillus fumigatus. Current diagnostic tests for the disease lack sensitivity as well as specificity, and culture of the fungus from invasive lung biopsy, considered the gold standard for IPA detection, is slow and often not possible in critically ill patients. In a previous study, we reported the development of a novel non-invasive procedure for IPA diagnosis based on antibody-guided positron emission tomography and magnetic resonance imaging (immunoPET/MRI) using a [64Cu]DOTA-labeled mouse monoclonal antibody (mAb), mJF5, specific to Aspergillus. To enable translation of the tracer to the clinical setting, we report here the development of a humanised version of the antibody (hJF5), and pre-clinical imaging of lung infection using a [64Cu]NODAGA-hJF5 tracer. The humanised antibody tracer shows a significant increase in in vivo biodistribution in A. fumigatus infected lungs compared to its radiolabeled murine counterpart [64Cu]NODAGA-mJF5. Using reverse genetics of the pathogen, we show that the antibody binds to the antigenic determinant 1,5-galactofuranose (Galf) present in a diagnostic mannoprotein antigen released by the pathogen during invasive growth in the lung. The absence of the epitope Galf in mammalian carbohydrates, coupled with the enhanced imaging capabilities of the hJF5 antibody, means that the [64Cu]NODAGA-hJF5 tracer developed here represents an ideal candidate for the diagnosis of IPA and translation to the clinical setting.This work was supported by the European Union Seventh Framework Programme FP7/2007-2013 under Grant 602820, the Deutsche Forschungsgemeinschaft (Grant WI3777/1-2 to SW), and the Werner Siemens Foundation. We thank Sven Krappman for use of the A. fumigatustdTomato strain, and acknowledge the Imaging Centre Essen (IMCES) for assistance with optical imaging of lungs

Trženjsko komuniciranje izdelkov višjega cenovnega razreda v majhnem podjetju

Invasive pulmonary aspergillosis (IPA) is a life-threatening lung disease caused by the fungus Aspergillus fumigatus, and is a leading cause of invasive fungal infection-related mortality and morbidity in patients with hematological malignancies and bone marrow transplants. We developed and tested a novel probe for noninvasive detection of A. fumigatus lung infection based on antibody-guided positron emission tomography and magnetic resonance (immunoPET/MR) imaging. Administration of a [(64)Cu]DOTA-labeled A. fumigatus-specific monoclonal antibody (mAb), JF5, to neutrophil-depleted A. fumigatus-infected mice allowed specific localization of lung infection when combined with PET. Optical imaging with a fluorochrome-labeled version of the mAb showed colocalization with invasive hyphae. The mAb-based newly developed PET tracer [(64)Cu]DOTA-JF5 distinguished IPA from bacterial lung infections and, in contrast to [(18)F]FDG-PET, discriminated IPA from a general increase in metabolic activity associated with lung inflammation. To our knowledge, this is the first time that antibody-guided in vivo imaging has been used for noninvasive diagnosis of a fungal lung disease (IPA) of humans, an approach with enormous potential for diagnosis of infectious diseases and with potential for clinical translation

Genetic Analysis of 15 Protein Folding Factors and Proteases of the Escherichia coli Cell Envelope

Each cell hosts thousands of proteins that vary greatly in abundance, structure, and chemical properties. To ensure that all proteins are biologically active and properly localized, efficient quality control systems have evolved. While the structure, function, and regulation of some individual protein folding factors and proteases were resolved up to atomic resolution, others remain poorly characterized. In addition, little is known about which factors are required for viability under specific stress conditions. We therefore determined the physiological implications of 15 factors of the E. coli cell envelope by an integrated genetic approach comprising phenotypic analyses. Our data indicate that surA and tsp null mutations are a lethal combination in rich medium, that surA dsbA and surA dsbC double mutants are temperature sensitive, and that surA ptrA, surA yfgC, dsbA fkpA, degP tsp, degP ppiD, tsp ppiD, and degP dsbA double mutants are temperature sensitive in rich medium containing 0.5 M NaCl, while degP dsbA, degP yfgC, tsp ydgD, and degP tsp double mutants do not grow in the presence of SDS/EDTA. Furthermore, we show that in degP dsbA, degP tsp, and degP yfgC double mutants a subpopulation of LamB exists as unfolded monomers. In addition, dsbA null mutants expressed lower levels of the outer membrane proteins LptD, LamB, FhuA, and OmpW while FhuA levels were reduced in surA single and degP ppiD double mutants. Lower FhuA levels in degP ppiD strains depend on Tsp, since in a tsp degP ppiD triple mutant FhuA levels are restored