Methods and compounds for the treatment of mucus hypersecretion

Abstract

A method of treating mucus hypersecretion, the causative factor in chronic obstructive pulmonary disease (COPD), asthma and other clinical conditions involving COPD, comprises administering a compound that inhibits exocytosis in mucus secreting cells or neurones that control or direct mucus secretion. Also described is a compound, for use in the treatment of hypersecretion of mucus, which inhibits mucus secretion by inhibiting mucus secretion by mucus secreting cells, and/or inhibiting neurotransmitter release from neuronal cells controlling or directing mucus secretion.

1 - 54 . (canceled) 
     
     
         55 . An agent for treating pain comprising a modified botulinum neurotoxin comprising a botulinum neurotoxin H N , a botulinum neurotoxin L chain, and no functional H C  domain, the modified botulinum neurotoxin being covalently coupled to substance P so that the modified botulinum neurotoxin no longer binds to botulinum neurotoxin receptors at a neuromuscular junction with the same affinity as the native botulinum neurotoxin. 
     
     
         56 . The agent of  claim 55 , wherein the botulinum neurotoxin is a botulinum toxin selected from the group consisting of serotype A, serotype B, serotype C 1 , serotype D, serotype E, serotype F and serotype G. 
     
     
         57 . The agent of  claim 55 , wherein the botulinum neurotoxin is botulinum toxin serotype A. 
     
     
         58 . The agent of  claim 55 , wherein the agent is a fusion protein. 
     
     
         59 . The agent of  claim 55 , wherein the L chain is obtained from a botulinum toxin selected from the group consisting of botulinum toxin serotype A, serotype B, serotype C 1 , serotype D, serotype E, serotype F and serotype G. 
     
     
         60 . The agent of  claim 55 , wherein the H N  is obtained from a botulinum toxin selected from the group consisting of botulinum toxin serotype A, serotype B, serotype C 1 , serotype D, serotype E, serotype F and serotype G. 
     
     
         61 . A plasmid encoding a modified clostridial neurotoxin, comprising: (a) a first nucleotide sequence comprising; (i) a first nucleotide segment encoding an amino acid sequence comprising substance P as the targeting moiety; and (ii) a second nucleotide segment encoding an amino acid sequence comprising a translocation domain (H N ) of a clostridial neurotoxin or a fragment thereof, which fragment translocates a light chain (L-chain) or L-chain fragment of a clostridial neurotoxin across an endosome membrane, wherein the first and second nucleotide segments encode an amino acid sequence comprising a fusion protein of a translocation domain and substance P, and wherein the H C  has been removed from said clostridial neurotoxin or has been modified so as to reduce the ability of the clostridial neurotoxin to bind to a receptor for the H C  at a neuromuscular junction; (b) a second nucleotide sequence encoding an amino acid sequence comprising the L-chain or an L-chain fragment of a clostridial neurotoxin, which L-chain fragment comprises the active protease domain of L-chain and (c) an element for plasmid replication by a host cell. 
     
     
         62 . A method of making a modified clostridial neurotoxin, the method comprising: (a) inserting the plasmid of  claim 61  into a suitable host cell, (b) culturing the host cell under conditions sufficient to express the clostridial neurotoxin, and (c) isolating the clostridial neurotoxin. 
     
     
         63 . A method of obtaining an agent for alleviating pain, the method comprising: (a) producing the plasmid of  claim 61  which encodes a modified clostridial neurotoxin; (b) inserting the plasmid into a suitable host cell; (c) culturing the host cell under conditions sufficient to express the clostridial neurotoxin; and isolating the clostridial neurotoxin as the agent for alleviating pain. 
     
     
         64 . The method of  claim 63 , wherein the modified clostridial neurotoxin further comprises at least one spacer component. 
     
     
         65 . The method of  claim 63 , wherein the clostridial neurotoxin comprises an H N  and an L chain. 
     
     
         66 . The method of  claim 65 , wherein the H N  has an amino acid sequence identical to the translocation domain of a clostridial neurotoxin from an organism selected from the group consisting of  Clostridial beratti, Clostridial butyricum, Clostridial botulinum , and  Clostridial tetani.    
     
     
         67 . The method of  claim 65 , wherein the L-chain has an amino acid sequence identical to the light chain of a clostridial neurotoxin from an organism selected from the group consisting of  Clostridial beratti, Clostridial butyricum, Clostridial botulinum , and  Clostridial tetani.    
     
     
         68 . The method of  claim 65 , wherein the H N  has an amino acid sequence identical to the translocation domain of a botulinum toxin selected from the group consisting of botulinum toxin serotype A, serotype B, serotype C 1 , serotype D, seretype E, serotype F, and serotype G. 
     
     
         69 . The method of  claim 65 , wherein the H N  has an amino acid sequence identical to the translocation domain of a botulinum toxin selected from the group consisting of botulinum toxin serotype A, serotype B, serotype C 1 , serotype D, serotype E, serotype F, and serotype G, and wherein the L-chain has an amino acid sequence identical to the light chain of a botulinum toxin selected from the group consisting of botulinum toxin serotype A, serotype B, serotype C 1 , serotype D, serotype E, serotype F, and serotype G. 
     
     
         70 . The method of  claim 65 , wherein the H N  has an amino acid sequence identical to the translocation domain of botulinum toxin serotype A, and wherein L-chain has an amino acid sequence identical to the light chain of botulinum toxin serotype A. 
     
     
         71 . A compound comprising a modified botulinum neurotoxin comprising a botulinum neurotoxin H N , a botulinum neurotoxin L chain, wherein said modified neurotoxin is modified so as to disable or remove its H C  portion, the modified botulinum neurotoxin being covalently linked to a Targeting Moiety (TM) so that the modified botulinum neurotoxin no longer binds to botulinum neurotoxin receptors at a neuromuscular junction with the same affinity as the native botulinum neurotoxin. 
     
     
         72 . The compound of  claim 71 , wherein the TM is substance P. 
     
     
         73 . A plasmid encoding a modified clostridial neurotoxin, comprising: (a) a first nucleotide sequence comprising; (i) a first nucleotide segment encoding an amino acid sequence as the targeting moiety (TM); and (ii) a second nucleotide segment encoding an amino acid sequence comprising a translocation domain (H N ) of a clostridial neurotoxin or a fragment thereof, which fragment translocates a light chain (L-chain) or L-chain fragment of a clostridial neurotoxin across an endosome membrane, wherein the first and second nucleotide segments encode an amino acid sequence comprising a fusion protein of a translocation domain and a TM, and wherein the H C  has been disabled or removed from said clostridial neurotoxin so as to reduce the ability of the clostridial neurotoxin to bind to a receptor for the H C  at a neuromuscular junction; (b) a second nucleotide sequence encoding an amino acid sequence comprising the L-chain or an L-chain fragment of a clostridial neurotoxin, which L-chain fragment comprises the active protease domain of L-chain and (c) an element for plasmid replication by a host cell. 
     
     
         74 . The plasmid of  claim 73 , wherein the TM is substance P. 
     
     
         75 . A method of making a modified clostridial neurotoxin, the method comprising: (a) inserting the plasmid of  claim 73  into a suitable host cell, (b) culturing the host cell under conditions sufficient to express the clostridial neurotoxin, and (c) isolating the clostridial neurotoxin. 
     
     
         76 . A method of obtaining a compound, the method comprising: (a) producing the plasmid of  claim 73  which encodes a modified clostridial neurotoxin; (b) inserting the r plasmid into a suitable host cell; (c) culturing the host cell under conditions sufficient to express the clostridial neurotoxin; and isolating the clostridial neurotoxin.