Delivering small molecules that bind botulinum toxin along with a monoclonal antibody (MAb) that binds them is being developed as a new strategy for rapidly and safely clearing this highly lethal toxin from the body, according to helminthologist Charles Shoemaker at the Tufts University Cummings School of Veterinary Medicine in North Grafton, Mass., and his collaborators there and at Thomas Jefferson University in Philadelphia, Pa. Their results from testing this strategy in mice appear in the February 2010 issue of Infection and Immunity (78:756-763).

Antidotes for biological toxins typically contain polyclonal antibodies, which are expensive to manufacture and stockpile, according to Shoemaker. In comparison, because his new approach is designed around specifically programmed, low-molecular-weight binding agents and a single, or universal, MAb, it offers a cheaper option with reduced overall development time, he says. The process could be used for making a variety of antitoxins with which to treat people exposed to one or more toxins as part of a bioterrorism attack or any from a broad range of natural toxins and also perhaps cytokines that the immune system releases during chronic illnesses.

To demonstrate the feasibility of their approach, Shoemaker and his collaborators derived single-chain Fv fragments (scFvs) from antibodies with a high affinity for serotype A botulinum toxin, which blocks transmission between nerves and muscles. The researchers tested six recombinant scFvs, each of which attaches to different sites along the botulinum toxin molecule. However, they also used a single anti-tag IgG1 MAb that recognizes all six scFvs.

Shoemaker and his team then tested which MAb-scFvs combinations could protect mice against botulinum toxin, administering doses that would be 1,000 to 10,000 times the 50% lethal dose. Untreated mice die within hours of receiving the toxin. However, mice treated by infusion with the MAb-scFvs mixture survive, and the toxin is excreted via the liver before it causes ill effects. Mice are not protected against botulinum toxin when they receive only scFvs or MAb without the other.

Although recombinant scFvs behaved well in these proof-of-principle experiments involving mice, alternative binding agents are being considered, including nanobodies and aptamers, according to Shoemaker. Since that study, they tried nanobodies as the binding agents and expressed different antitoxin nanobodies as a single protein agent. "With just a single agent, we can attract multiple MAbs to the toxin," he says.

The Tufts researchers also are developing binding agents that target other types of botulinum toxin, Shiga toxin, and toxins from Clostridium difficile. Additionally, they plan to develop binding agents for clearing cytokines that play a part in causing inflammation responses that injure the host and in autoimmune diseases. Showing that the method is flexible and not limited to botulism "could make commercialization of the approach more appealing," Shoemaker says.

"There's strong value in this idea, and it looks like it can be generalized for a lot of different toxins or infectious agents," says Kim Janda, a chemical biologist at Scripps Research Institute in La Jolla, Calif. He hopes that someone will evaluate whether this approach is safe for human patients. "It has all the right bells and whistles to make it worthy of clinical investigation," he says.

Carol Potera
Carol Potera is a freelance writer in Great Falls, Mont.