Recombinant Neurotoxin Derivatives Customized for Specific Indications
Background and Description of Technology:
Botulinum neurotoxin (BoNT) is one of the most poisonous substances known, with an LD50 of approximately 1ng per kg, and the ability to intoxicate via trans-epithelial routes. Because BoNT can be weaponized, it is considered a priority of the NIAID Biodefense Research Agenda. Significant government and private funds are currently being devoted to the developments of vaccines and antidotes that can protect civilian and military populations against BoNT poisoning. The current antitoxin is scarce, difficult to administer and can have life-threatening side effects. BoNT also has important therapeutic indications. Small doses precisely delivered to specific muscles have been used to treat diverse neuromuscular pathologies (e.g. dystonia, migraine, cerebral palsy) and more recently as a medical aesthetic product to temporarily eliminate facial wrinkles. One marketed brand, Botox, achieved sales of more than $1B in 2008.
A genetic engineering platform has been developed to manipulate and express Clostridial neurotoxin (NT) genes. It is based on novel compositions and methods that enable the production of recombinant, full-length physiologically active Clostridial neurotoxin derivatives that retain the native conformation and disulfide bonding of the natural toxins. These methods provide a platform for producing NT derivatives customized to specific therapeutic indications. They enable the NT derivatives to mimic the trafficking route and neuronal targeting of natural clostridial NTs, and therefore provide important advantages over other available methods. The expression systems utilized enable high- level production of the NT derivatives, and facilitates their efficient purification. Work thus far has concentrated on the NT produced by Clostridium Botulinum serotype A (BoNT/A). BoNT/A derivatives have been produced in enzymatically active and inactive forms. The active form is similar in potency to pharmaceutical preparations of Botulinum Neurotoxin A (BoNT A) with respect to systemic toxicity, and in the standard mouse phrenic nerve hemidiaphragm assay. The enzymatically inactive form has significantly reduced systemic toxicity, and is being used to develop antidotes and vaccines to BoNT poisoning. Chimeric BoNT/A-GFP (Green Fluorescent Protein) derivatives have been developed to evaluate systemic and intra- cellular BoNT/A trafficking. Additional chimeric BoNT derivatives are being developed to deliver specific agents to neurons via the neuromuscular junction, including BoNT antagonists designed to rescue BoNT poisoned neurons.
Potential applications include:
BoNT Therapeutics: The technology developed provides the first recombinant system capable of producing BoNT/A derivatives with similar potency to pharmaceutical BoNT/A preparations. It enables the manipulation of the BoNT genes via mutations and rearrangements that are not possible to perform in Clostridium cultures. It therefore facilitates the introduction of pharmaceutically useful modifications that can provide competitive advantages in BoNT therapeutics for a variety of indications: Aesthetic (rhytides), gastroenterologic, genitourinary and neurologic (muscle spasticity and dystonias).
Biodefense: The NT derivatives described have definitive advantages for vaccine and antidote development. Because they can be rendered non-toxic by point mutations targeting the enzymatic and substrate-binding activities responsible for toxicity, the removal of toxicity can be accomplished with minimal structural alteration. This is expected to improve the effectiveness of vaccines, and allow their delivery by trans- epithelial (e.g. inhalation) routes of administration. The platform methodology is also being exploited to develop BoNT antidotes that will specifically traffic to and target BoNT- affected neurons. Antidotes that can be effective after BoNT-induced paralysis has set in provide unique practical advantages, and may be capable of rescuing BoNT- affected individuals that do not have access to long-term maintenance on respirators.
Neuronal Targeting Platform: The ability of the recombinant BoNT products to specifically target neurons at the neuromuscular junction makes it possible to use this system to deliver therapeutic agents directly to neurons. The neuronal targeting technology can be extended to agents requiring delivery to the central nervous system, using the trafficking properties of the related Tetanus neurotoxins (TeNT, also produced by Clostridium species and with extensive homology to BoNTs).
A patent application has been filed on this technology and NYU is seeking commercial partners to develop therapeutics for the various areas described above.