Abstract
Gastrointestinal nematodes (GINs) of humans, e.g., hookworms, negatively impact childhood growth, cognition, nutrition, educational attainment, income, productivity, and pregnancy. Hundreds of millions of people are targeted with mass drug administration (MDA) of donated benzimidazole anthelmintics. However, benzimidazole efficacy against GINs is suboptimal, and reduced/low efficacy has been seen. Developing an anthelmintic for human MDA is daunting: it must be safe, effective, inexpensive, stable without a cold chain, and massively scalable. Bacillus thuringiensis crystal protein 5B (Cry5B) has anthelmintic properties that could fill this void. Here, we developed an active pharmaceutical ingredient (API) containing B. thuringiensis Cry5B compatible with MDA. We expressed Cry5B in asporogenous B. thuringiensis during vegetative phase, forming cytosolic crystals. These bacteria with cytosolic crystals (BaCC) were rendered inviable (inactivated BaCC [IBaCC]) with food-grade essential oils. IBaCC potency was validated in vitro against nematodes. IBaCC was also potent in vivo against human hookworm infections in hamsters. IBaCC production was successfully scaled to 350 liters at a contract manufacturing facility. A simple fit-for-purpose formulation to protect against stomach digestion and powdered IBaCC were successfully made and used against GINs in hamsters and mice. A pilot histopathology study and blood chemistry workup showed that five daily consecutive doses of 200 mg/kg body weight Cry5B IBaCC (the curative single dose is 40 mg/kg) was nontoxic to hamsters and completely safe. IBaCC is a safe, inexpensive, highly effective, easy-to-manufacture, and scalable anthelmintic that is practical for MDA and represents a new paradigm for treating human GINs.
Highlights
Among the neglected tropical diseases, soil-transmitted helminths/nematodes (STHs/STNs) or gastrointestinal nematodes (GINs) collectively affect the largest number of people, with a global estimate of .1.5 billion infected individuals [1]
We reasoned that delivering crystal protein 5B (Cry5B) in a live bacterium (B. thuringiensis or otherwise) to humans would be problematic given
Paraprobiotic Cure for Intestinal Nematodes that (i) B. thuringiensis is closely related to Bacillus cereus, which can cause food poisoning, (ii) release of live recombinant bacteria has environmental concerns, because live bacteria in the soil could select for Cry5B resistance against free-living stages of hookworms, (iii) live bacteria could replicate in the human gastrointestinal tract, (iv) there are uncertainties and significant safety concerns in the responses of billions of people to live bacteria, (v) degradation of live bacteria during storage could reduce potency and stability, and (vi) delivery of a stable live bacterial therapeutic around the world for mass drug administration would be difficult to achieve [36,37,38,39,40]
Summary
Among the neglected tropical diseases, soil-transmitted helminths/nematodes (STHs/STNs) or gastrointestinal nematodes (GINs) collectively affect the largest number of people, with a global estimate of .1.5 billion infected individuals [1]. Developing 3D-Cry proteins as large-scale ingested therapeutics compatible with human mass drug administration, requires a very different set of considerations than application as topical or transgenic biopesticides and insecticides. We address these considerations, developing an active pharmaceutical ingredient (API) based on Cry5B compatible with safety, cost, scale, ease of production, and stability required for human mass drug administration
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