Development of a 13C Stable Isotope Assay for Dipeptidyl Peptidase-4 Enzyme Activity A New Breath Test for Dipeptidyl Peptidase Activity

Abstract

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 13C-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 13C-tripeptide was added to cells in the presence and absence of the DPP4 inhibitor Sitagliptin. Gas samples were collected from the cell headspace and 13CO2 content quantified by isotope ratio mass spectrometry (IRMS). DPP4 was highly expressed in Caco-2 cells compared to HeLa cells and using the 13C-tripeptide, we detected a high 13CO2 signal from Caco2 cells. Addition of Sitaglitpin to Caco2 cells significantly inhibited this 13CO2 signal. 13C-assay DPP4 activity correlated positively with the enzyme activity detected using a colorimetric substrate. We have developed a selective, non-invasive, 13C-assay for DPP4 that could have broad translational applications in diabetes and gastrointestinal disease.

The perceptions of natural compounds against dipeptidyl peptidase 4 in diabetes: fromin silicotoin vivo

“…108 Yazbeck et al have derived a new DPP4 substrate with 13 C isotope that can be released upon reaction with DPP-4. 111 As such, the requirement to homogenize the biopsy is completely exonerated, leading to higher correlation as compared with its conventional counterparts. 111 Myocytes and pancreatic cells are often used in the cell-based assays to discover DPP-4 inhibitors.…”

Circulating Dipeptidyl Peptidase Activity Is a Potential Biomarker for Inflammatory Bowel Disease

“…DPP4 enzyme activity and expression have been investigated as a potential disease biomarker for a range of diseases, including IBD ( 15 , 16 ). Xiao et al ( 17 ) reported decreased circulating DPP (cDPP) activity in patients with IBD compared with healthy patients.…”