Optimized LC-MS/MS Method for the Detection of ppCCK(21–44): A Surrogate to Monitor Human Cholecystokinin Secretion

The hormone cholecystokinin (CCK) is secreted postprandially
from
duodenal enteroendocrine cells and circulates in the low picomolar
range. Detection of this digestion and appetite-regulating hormone
currently relies on the use of immunoassays, many of which suffer
from insufficient sensitivity in the physiological range and cross-reactivity
problems with gastrin, which circulates at higher plasma concentrations.
As an alternative to existing techniques, a liquid chromatography
and mass spectrometry-based method was developed to measure CCK-derived
peptides in cell culture supernatants. The method was initially applied
to organoid studies and was capable of detecting both CCK8 and an
N-terminal peptide fragment (prepro) ppCCK(21–44) in supernatants
following stimulation. Extraction optimization was performed using
statistical modeling software, enabling a quantitative LC-MS/MS method
for ppCCK(21–44) capable of detecting this peptide in the low
pM range in human plasma and secretion buffer solutions. Plasma samples
from healthy individuals receiving a standardized meal (Ensure) after
an overnight fast were analyzed; however, the method only had sensitivity
to detect ppCCK(21–44). Secretion studies employing human intestinal
organoids and meal studies in healthy volunteers confirmed that ppCCK(21–44)
is a suitable surrogate analyte for measuring the release of CCK in
vitro and in vivo.