Peripheral and Mucosal B Cells From Celiac Disease Patients Show Increased Expression of CXCR3 and IgA Switch Markers

ponding to subdivisions of the 33mer based on the ability of ALV001 to cut several times within this gluten epitope. RESULTS: Mass spectrometry detected 33mer and all five peptide fragments resulting from degradation of the 33mer by ALV001. A large linear range was observed, with detection as low as 1 ng/ml of the peptides of interest. Intraday coefficient of variability was typically less than 5%. ALV001 cut within the 33mer epitope in the context of wheat gluten, releasing all five fragments that initially rose in concentration but plateaued as ALV001-mediated degradation slows. Using a clinical dose (150 mg, 0.3 mg/ml), ALV001 degraded 98% (10 minutes) and 99.5% (30 minutes) of the 33mer epitope. ALV001 and ALV002 functioned cooperatively; ALV002 degraded the five relatively stable peptide fragments as ALV001 activity plateaued. SUMMARY: A mass spectrometry assay was established to quantify the ability of ALV003 to degrade 33mer within wheat bread gluten in the context of a complex meal. Using this methodology, it was possible to distinguish separate but complementary contributions of ALV001 and ALV002 to 33mer degradation. The data support a model for ALV003-mediated gluten degradation in which ALV001 rapidly cleaves gluten adjacent to glutamine residues, releasing small proline-rich peptide fragments that serve as substrates for the prolyl endopeptidase ALV002. Data from a simulated gastric environment suggest that ALV003 should degrade ingested gluten in patients with celiac disease.