Enzyme treatment of foodstuffs for celiac sprue

Abstract

Administering an effective dose of glutenase to a Celiac or dermatitis herpetiformis patient reduces levels of toxic gluten oligopeptides, thereby attenuating or eliminating the damaging effects of gluten.

What is claimed is: 
     
         1 . A method of treating Celiac Sprue and/or dermatitis herpetiformis, the method comprising:
 administering to a patient an effective dose of a glutenase;   wherein said glutenase attenuates gluten toxicity in said patient.   
     
     
         2 . The method according to  claim 1 , wherein said glutenase is an enzyme capable of cleaving a pretreated substrate, wherein said pretreated substrate comprises one or more gliadin, hordein, secalin or avenin proteins after pretreatment with physiological quantities of gastric and pancreatic proteases, and wherein said glutenase, when added to a reaction mixture comprising said pretreated substrate, increases the concentration of free NH 2 -termini by at least about 25%, and/or reduces the residual molar concentration of oligopeptides greater than about 1000 Da by at least about 2-fold, and/or reduces the potency by which said pretreated substrate antagonizes binding of (SEQ ID NO:17) PQPELPYPQPQLP to HLA-DQ2. 
     
     
         3 . The method according to  claim 1 , wherein said glutenase is an enzyme having at least about 20% sequence identity at the amino acid level to one of: prolyl endopeptidase from  F. meningosepticum , DCP I from  E. coli  and DPP IV from  Aspergillus fumigatus  (Genbank accession number U87950). 
     
     
         4 . The method according to  claim 1 , wherein said glutenase has a specific activity of at least 2.5 U/mg for cleavage of a peptide comprising one of more motifs selected from the group consisting of Gly-Pro-pNA, Z-Gly-Pro-pNA, and Hip-His-Leu and/or a kcat/Km of at least about 2.5 s −1  M −1  for cleavage of a peptide selected from the group consisting of (SEQ ID NO:1) QLQPFPQPQLPY, (SEQ ID NO:3) PQPQLPYPQPQLPY, (SEQ ID NO:13) QPQQSFPQQQ, (SEQ ID NO:14) QLQPFPQPELPY, (SEQ ID NO:15) PQPELPYPQPELPY, and (SEQ ID NO:16) QPQQSFPEQQ. 
     
     
         5 . The method according to  claim 1 , wherein said glutenase is an enzyme belonging to the classification group EC 3.4.21.26, EC 3.4.14.5, or EC 3.4.15.1. 
     
     
         6 . The method according to  claim 1 , wherein said glutenase is formulated with a pharmaceutically acceptable excipient. 
     
     
         7 . The method according to  claim 1 , wherein said glutenase is admixed with food. 
     
     
         8 . The method according to  claim 1 , wherein said glutenase is stable to acid conditions. 
     
     
         9 . The method according to  claim 1 , wherein said glutenase is administered orally. 
     
     
         10 . The method according to  claim 9 , wherein said glutenase is contained in a formulation that comprises an enteric coating. 
     
     
         11 . A formulation for use in treatment of Celiac Sprue and/or dermatitis herpetiformis, comprising:
 an effective dose of glutenase and a pharmaceutically acceptable excipient.   
     
     
         12 . The formulation according to  claim 11 , wherein said glutenase is an enzyme capable of cleaving a pretreated substrate, wherein said pretreated substrate comprises one or more gliadin, hordein, secalin or avenin proteins after pretreatment with physiological quantities of gastric and pancreatic proteases, wherein said glutenase, when added to a reaction mixture comprising said pretreated substrate, increases the concentration of free NH 2 -termini by at least about 25%, and/or reduces the residual molar concentration of oligopeptides greater than about 1000 Da by at least about 2-fold, and/or reduces the potency by which said pretreated substrate antagonizes binding of (SEQ ID NO:17) PQPELPYPQPQLP to HLA-DQ2. 
     
     
         13 . The formulation according to  claim 12 , wherein said glutenase is an enzyme having at least about 20% sequence identity at the amino acid level to one of: prolyl endopeptidase from  F. meningosepticum , DCP I from rabbit and DPP IV from  Aspergillus fumigatus  (Genbank accession number U87950). 
     
     
         14 . The formulation according to  claim 12 , wherein said glutenase has a specific activity of at least 2.5 U/mg for cleavage of a peptide comprising one of more motifs selected from the group consisting of Gly-Pro-pNA, Z-Gly-Pro-pNA, and Hip-His-Leu and/or a kcat/Km of at least about 2.5 s −1  M −1  for cleavage of a peptide selected from the group consisting of (SEQ ID NO:1) QLQPFPQPQLPY, (SEQ ID NO:3) PQPQLPYPQPQLPY, (SEQ ID NO:13) QPQQSFPQQQ, (SEQ ID NO:14) QLQPFPQPELPY, (SEQ ID NO:15) PQPELPYPQPELPY, and (SEQ ID NO:16) QPQQSFPEQQ. 
     
     
         15 . The formulation according to  claim 12 , wherein said glutenase is an enzyme belonging to the classification group EC 3.4.21.26, EC 3.4.14.5, or EC 3.4.15.1. 
     
     
         16 . The formulation according to  claim 12 , wherein said glutenase is stable to acid conditions. 
     
     
         17 . The formulation according to  claim 12 , wherein said formulation is suitable for oral administration. 
     
     
         18 . The formulation according to  claim 12 , wherein said formulation comprises an enteric coating. 
     
     
         19 . A method for treating a foodstuff to render said foodstuff less toxic to a Celiac Sprue patient, said method comprising contacting said foodstuff with a glutenase. 
     
     
         20 . The method of  claim 19 , wherein said glutenase is prolyl endopeptidase. 
     
     
         21 . A purified oligopeptide selected from the group consisting of (SEQ ID NO:12) LQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPF, a deamidated counterpart oligonucleotide, an analog oligopeptide, and a conjugate of tissue transgluatonimase and a counterpart oligonucleotide. 
     
     
         22 . A method of diagnosing Celiac Sprue in an individual said method comprising determining the presence of T cells in said individual that are reactive towards to the oligopeptide of  claim 21 . 
     
     
         23 . A method of diagnosing Celiac Sprue in an individual said method comprising determining the presence in said individual of antibodies specific for the oligopeptide of  claim 21 , or a derivative or conjugate thereof. 
     
     
         24 . A method for diagnosing Celiac Sprue that comprises detecting the presence of an oligopeptide of  claim 21  in a tissue, bodily fluid, or stool of an individual.