It is known that many chemotherapeutics induce cellular apoptosis over hours to days. During apoptosis, numerous cellular
proteases are activated, most canonically the
caspases. We speculated that detection of proteolytic fragments released from apoptotic cells into the peripheral blood may serve as a unique
indicator of
chemotherapy-induced cell death. Here we used an enzymatic labeling process to positively enrich free
peptide α-
amines in the plasma of
hematologic malignancy patients soon after beginning treatment. This N-terminomic approach largely avoids interference by high-abundance
proteins that complicate traditional plasma proteomic analyses. Significantly, by mass spectrometry methods, we found strong
biological signatures of apoptosis directly in the postchemotherapy plasma, including numerous
caspase-cleaved
peptides as well as relevant
peptides from apoptotic and cell-
stress proteins second mitochondria-derived activator of
caspases,
HtrA serine peptidase 2, and
activating transcription factor 6. We also treated hematologic
cancer cell lines with clinically relevant chemotherapeutics and monitored proteolytic fragments released into the media. Remarkably, many of these
peptides coincided with those found in patient samples. Overall, we identified 153 proteolytic
peptides in postchemotherapy patient plasma as potential indicators of cellular apoptosis. Through targeted quantitative proteomics, we verified that many of these
peptides were indeed increased post- vs. prechemotherapy in additional patients. Our findings reveal that numerous proteolytic fragments are released from dying
tumor cells. Monitoring posttreatment proteolysis may lead to a novel class of inexpensive, rapid
biomarkers of cell death.