Novel botulinum neurotoxin and its derivatives

Abstract

Provided herein are Clostridial Botulinum neurotoxin (BoNT) polypeptides of a novel serotype (BoNT/X) and methods of making and using the BoNT polypeptides, e.g., in therapeutic applications.

What is claimed is: 
     
         1 .- 175 . (canceled) 
     
     
         176 . A chimeric Clostridial  Botulinum  neurotoxin (BoNT) polypeptide comprising:
 (a) a protease domain;   (b) a linker region;   (c) a translocation domain; and   (d) a receptor binding domain,   wherein (a) and (c) are from BoNT serotype X, and wherein (d) is not from BoNT serotype X.   
     
     
         177 . The chimeric BoNT polypeptide of  claim 176 , wherein the receptor binding domain is modified. 
     
     
         178 . The chimeric BoNT polypeptide of  claim 177 , wherein the receptor binding domain comprises a substitution mutation corresponding to C1240S or C1240A in SEQ ID NO: 1. 
     
     
         179 . The chimeric BoNT polypeptide of  claim 176 , wherein the receptor binding domain is from serotype selected from the group consisting of A, B, C, D, E, F, and G. 
     
     
         180 . The chimeric BoNT polypeptide of  claim 176  comprising an amino acid sequence that has at least 85% identity to any one of SEQ ID NOs: 22-24, wherein the polypeptide does not have the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2. 
     
     
         181 . The chimeric BoNT polypeptide of  claim 180 , comprising an amino acid sequence of any one of SEQ ID NOs: 22-24. 
     
     
         182 . The chimeric BoNT polypeptide of  claim 176 , wherein the linker region comprises an artificial linker. 
     
     
         183 . The chimeric BoNT polypeptide of  claim 182 , wherein the artificial linker comprises a cleavage site of a protease. 
     
     
         184 . The chimeric BoNT polypeptide of  claim 183 , wherein the protease is selected from the group consisting of thrombin, TEV, PreScission (3C protease), Factor Xa, Enterokinase, and SUMO protease. 
     
     
         185 . The chimeric BoNT polypeptide of  claim 184 , wherein the linker comprises an amino acid sequence of any of SEQ ID NOs: 50-56. 
     
     
         186 . The chimeric BoNT polypeptide of  claim 176 , comprising one or more substitution mutation(s) in a position corresponding to R360, Y363, H227, E228, or H231 in SEQ ID NO: 1. 
     
     
         187 . The chimeric BoNT polypeptide of  claim 186 , wherein the one or more substitution mutation(s) corresponds to R360A/Y363F, H227Y, E228Q, or H231Y in SEQ ID NO: 1. 
     
     
         188 . A chimeric molecule comprising a first portion linked to a second portion, wherein the first portion is a chimeric BoNT polypeptide of  claim 186 . 
     
     
         189 . The chimeric molecule of  claim 188 , wherein the first portion and the second portion are linked covalently. 
     
     
         190 . The chimeric molecule of  claim 189 , wherein the first portion and the second portion are linked non-covalently. 
     
     
         191 . The chimeric molecule of  claim 189 , wherein the second portion is selected from the group consisting of a small molecule, a nucleic acid, a short polypeptide and a protein. 
     
     
         192 . The chimeric molecule of  claim 191 , wherein the second portion is a bioactive molecule. 
     
     
         193 . The chimeric molecule of  claim 191 , wherein the second portion is a non-polypeptide drug. 
     
     
         194 . A method of treating a condition associated with overactive neurons or glands, the method comprising administering a therapeutically effective amount of the modified BoNT polypeptide of  claim 176 . 
     
     
         195 . The method of  claim 194 , wherein the condition is selected from the group consisting of: spasmodic dysphonia, spasmodic torticollis, laryngeal dystonia, oromandibular dysphonia, lingual dystonia, cervical dystonia, focal hand dystonia, blepharospasm, strabismus, hemifacial spasm, eyelid disorder, cerebral palsy, focal spasticity and other voice disorders, spasmodic colitis, neurogenic bladder, anismus, limb spasticity, tics, tremors, bruxism, anal fissure, achalasia, dysphagia and other muscle tone disorders and other disorders characterized by involuntary movements of muscle groups, lacrimation, hyperhydrosis, excessive salivation, excessive gastrointestinal secretions, secretory disorders, pain from muscle spasms, headache pain, dermatological or aesthetic/cosmetic conditions, obesity/reduced appetite.