Isomerase Therapeutics Ltd.

Streptomyces iranensisHM 35 is an alternative rapamycin producer toStreptomyces rapamycinicus. Targeted genetic modification of rapamycin-producing actinomycetes is a powerful tool for the directed production of rapamycin derivatives, and it has also revealed some key features of the molecular biology of rapamycin formation inS. rapamycinicus.The approach depends upon efficient conjugational plasmid transfer fromEscherichia colitoStreptomyces, and the failure of this step has frustrated its application toStreptomyces iranensisHM 35. Here, by systematically optimizing the process of conjugational plasmid transfer, including screening of various media, and by defining optimal temperatures and concentrations of antibiotics and Ca2+ions in the conjugation media, we have achieved exconjugant formation for each of a series of gene deletions inS. iranensisHM 35. Among them wererapK, which generates the starter unit for rapamycin biosynthesis, andhutF, encoding a histidine catabolizing enzyme. The protocol that we have developed may allow efficient generation of targeted gene knockout mutants ofStreptomycesspecies that are genetically difficult to manipulate.IMPORTANCEThe developed protocol of conjugational plasmid transfer fromEscherichia colitoStreptomyces iranensismay allow efficient generation of targeted gene knockout mutants of other genetically difficult to manipulate, but valuable,Streptomycesspecies.

Inhibition of host-encoded targets, such as the cyclophilins, provides an opportunity to generate potent high barrier to resistance antivirals for the treatment of a broad range of viral diseases. However, many host-targeted agents are natural products, which can be difficult to optimize using synthetic chemistry alone. We describe the orthogonal combination of bioengineering and semisynthetic chemistry to optimize the drug-like properties of sanglifehrin A, a known cyclophilin inhibitor of mixed nonribosomal peptide/polyketide origin, to generate the drug candidate NVP018 (formerly BC556). NVP018 is a potent inhibitor of hepatitis B virus, hepatitis C virus (HCV), and HIV-1 replication, shows minimal inhibition of major drug transporters, and has a high barrier to generation of both HCV and HIV-1 resistance.

Malaria remains a major global health problem. Emerging resistance to existing antimalarial drugs drives the search for new antimalarials, and protein translation is a promising pathway to target. Here we explore the potential of the aminoacyl-tRNA synthetase (ARS) family as a source of antimalarial drug targets. First, a battery of known and novel ARS inhibitors was tested against Plasmodium falciparum cultures, and their activities were compared. Borrelidin, a natural inhibitor of threonyl-tRNA synthetase (ThrRS), stands out for its potent antimalarial effect. However, it also inhibits human ThrRS and is highly toxic to human cells. To circumvent this problem, we tested a library of bioengineered and semisynthetic borrelidin analogs for their antimalarial activity and toxicity. We found that some analogs effectively lose their toxicity against human cells while retaining a potent antiparasitic activity both in vitro and in vivo and cleared malaria from Plasmodium yoelii-infected mice, resulting in 100% mice survival rates. Our work identifies borrelidin analogs as potent, selective, and unexplored scaffolds that efficiently clear malaria both in vitro and in vivo.