Abstract
Rabies is a neglected zoonotic disease that has no effective treatment after onset of illness. However the disease can be prevented effectively by prompt administration of post exposure prophylaxis which includes administration of passive immunizing antibodies (Rabies Immune Globulin, RIG). Currently, human RIG suffers from many restrictions including limited availability, batch-to batch inconsistencies and potential for contamination with blood-borne pathogens. Anti-rabies monoclonal antibodies (mAbs) have been identified as a promising alternative to RIG. Here, we applied a plant-based transient expression system to achieve rapid, high level production and efficacy of the two highly potent anti-rabies mAbs E559 and 62-71-3. Expression levels of up to 490 mg/kg of recombinant mAbs were obtained in Nicotiana benthamiana glycosylation mutants by using a viral based transient expression system. The plant-made E559 and 62-71-3, carrying human-type fucose-free N-glycans, assembled properly and were structurally sound as determined by mass spectrometry and calorimetric density measurements. Both mAbs efficiently neutralised diverse rabies virus variants in vitro. Importantly, E559 and 62-71-3 exhibited enhanced protection against rabies virus compared to human RIG in a hamster model post-exposure challenge trial. Collectively, our results provide the basis for the development of a multi-mAb based alternative to RIG.
Highlights
Rabies is a zoonotic viral disease that continues to have no effective treatment after onset of symptoms
The current study describes the plant-based recombinant expression, purification, structural and functional characterisation of humanized anti rabies monoclonal antibodies (mAbs) E559 and 62-71-3. mAbs were expressed in ΔXT/FT, a Nicotiana benthamiana mutant supporting the synthesis of glycan-optimized fucose-free mAbs [10]
At 28 dpi, 33% of animals treated with mAb 62-71-3 and 20% of animals treated with E559 survived; while all animals treated with HRIG died
Summary
Rabies is a zoonotic viral disease that continues to have no effective treatment after onset of symptoms. Afucosylated therapeutic anti-cancer antibodies can exhibit superior in vitro and in vivo efficacy [18, 19] For these reasons, the anti-rabies mAbs were expressed in the ΔXTFT Nicotiana. Doi:10.1371/journal.pone.0159313.g007 because in spite of their common structure, individual mAbs often have unique and unpredictable responses to their environment related to their stability [23] This will be the case with the envisaged E559 / 62-71-3 cocktail where the data suggest E559 is less thermostable than 6271-3. As these antibodies have different binding sites, they provide the capacity to simultaneously bind RVG Use of these two Abs in a cocktail will enhance the neutralization of wider spectra of rabies viruses and reduce the chances of virus escape [9].
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