Characterization of a new Leishmania major strain for use in a controlled human infection model.

Leishmaniasis is widely regarded as a vaccine-preventable disease, but the costs required to reach pivotal Phase 3 studies and uncertainty about which candidate vaccines should be progressed into human studies significantly limits progress in vaccine development for this neglected tropical disease. Controlled human infection models (CHIMs) provide a pathway for accelerating vaccine development and to more fully understand disease pathogenesis and correlates of protection. Here, we describe the isolation, characterization and GMP manufacture of a new clinical strain of Leishmania major. Two fresh strains of L. major from Israel were initially compared by genome sequencing, in vivo infectivity and drug sensitivity in mice, and development and transmission competence in sand flies, allowing one to be selected for GMP production. This study addresses a major roadblock in the development of vaccines for leishmaniasis, providing a key resource for CHIM studies of sand fly transmitted cutaneous leishmaniasis.

ß-Glucan-Induced Trained Immunity in Dogs.

Several observations in the world of comparative immunology in plants, insects, fish and eventually mammals lead to the discovery of trained immunity in the early 2010's. The first demonstrations provided evidence that innate immune cells were capable of developing memory after a first encounter with some pathogens. Trained immunity in mammals was initially described in monocytes with the Bacille Calmette-Guerin vaccine (BCG) or prototypical agonists like ß-glucans. This phenomenon relies on epigenetic and metabolic modifications leading to an enhanced secretion of inflammatory cytokines when the host encounters homologous or heterologous pathogens. The objective of our research was to investigate the trained immunity, well-described in mouse and human, in other species of veterinary importance. For this purpose, we adapted an in vitro model of trained innate immunity in dogs. Blood enriched monocytes were stimulated with ß-glucans and we confirmed that it induced an increased production of pro-inflammatory and anti-microbial compounds in response to bacterial stimuli. These results constitute the first demonstration of trained immunity in dogs and confirm its signatures in other mammalian species, with an implication of cellular mechanisms similar to those described in mice and humans regarding cellular epigenetics and metabolic regulations.

Cryo-EM structure of pleconaril-resistant rhinovirus-B5 complexed to the antiviral OBR-5-340 reveals unexpected binding site.

Viral inhibitors, such as pleconaril and vapendavir, target conserved regions in the capsids of rhinoviruses (RVs) and enteroviruses (EVs) by binding to a hydrophobic pocket in viral capsid protein 1 (VP1). In resistant RVs and EVs, bulky residues in this pocket prevent their binding. However, recently developed pyrazolopyrimidines inhibit pleconaril-resistant RVs and EVs, and computational modeling has suggested that they also bind to the hydrophobic pocket in VP1. We studied the mechanism of inhibition of pleconaril-resistant RVs using RV-B5 (1 of the 7 naturally pleconaril-resistant rhinoviruses) and OBR-5-340, a bioavailable pyrazolopyrimidine with proven in vivo activity, and determined the 3D-structure of the protein-ligand complex to 3.6 Å with cryoelectron microscopy. Our data indicate that, similar to other capsid binders, OBR-5-340 induces thermostability and inhibits viral adsorption and uncoating. However, we found that OBR-5-340 attaches closer to the entrance of the pocket than most other capsid binders, whose viral complexes have been studied so far, showing only marginal overlaps of the attachment sites. Comparing the experimentally determined 3D structure with the control, RV-B5 incubated with solvent only and determined to 3.2 Å, revealed no gross conformational changes upon OBR-5-340 binding. The pocket of the naturally OBR-5-340-resistant RV-A89 likewise incubated with OBR-5-340 and solved to 2.9 Å was empty. Pyrazolopyrimidines have a rigid molecular scaffold and may thus be less affected by a loss of entropy upon binding. They interact with less-conserved regions than known capsid binders. Overall, pyrazolopyrimidines could be more suitable for the development of new, broadly active inhibitors.