Development of a C Stable Isotope Assay for Dipeptidyl Peptidase-4 Enzyme Activity A New Breath Test for Dipeptidyl Peptidase Activity.

Dipeptidyl peptidase-4 inhibitors (DPP4i) are a class of orally available, small molecule inhibitors for the management of Type-II diabetes. A rapid, real-time, functional breath test for DPP4 enzyme activity could help to define DPP4i efficacy in patients that are refractory to treatment. We aimed to develop a selective, non-invasive, stable-isotope C-breath test for DPP4. In vitro experiments were performed using high (Caco-2) and low (HeLa) DPP4 expressing cells. DPP gene expression was determined in cell lines by qRT-PCR. A DPP4 selective C-tripeptide was added to cells in the presence and absence of the DPP4 inhibitor Sitagliptin. Gas samples were collected from the cell headspace and CO content quantified by isotope ratio mass spectrometry (IRMS). DPP4 was highly expressed in Caco-2 cells compared to HeLa cells and using the C-tripeptide, we detected a high CO signal from Caco2 cells. Addition of Sitaglitpin to Caco2 cells significantly inhibited this CO signal. C-assay DPP4 activity correlated positively with the enzyme activity detected using a colorimetric substrate. We have developed a selective, non-invasive, C-assay for DPP4 that could have broad translational applications in diabetes and gastrointestinal disease.