Differentiation between Cell Death Modes Using Measurements of Different Soluble Forms of Extracellular Cytokeratin 18

Abstract

Cytokeratins are released from carcinoma cells by unclear mechanisms and are commonly used serum tumor markers (TPA, TPS, and CYFRA 21-1). We here report that soluble cytokeratin-18 (CK18) is released from human carcinoma cells during cell death. During necrosis, the cytosolic pool of soluble CK18 was released, whereas apoptosis was associated with significant release of caspase-cleaved CK18 fragments. These results suggested that assessments of different forms of CK18 in patient sera could be used to examine cell death modes. Therefore, CK18 was measured in local venous blood collected during operation of patients with endometrial tumors. In most patient sera, caspase-cleaved fragments constituted a minor fraction of total CK18, suggesting that tumor apoptosis is not the main mechanism for generation of circulating CK18. Monitoring of different CK18 forms in peripheral blood during chemotherapy of prostate cancer patients showed individual differences in the patterns of release. Importantly, several examples were observed where the increase of apoptosis-specific caspase-cleaved CK18 fragments constituted only a minor fraction of the total increase. These results suggest that cell death of epithelially derived tumors can be assessed in patient serum and suggest that tumor apoptosis may not necessarily be the dominating death mode in many tumors in vivo.

Refining Liver Safety Risk Assessment: Application of Mechanistic Modeling and Serum Biomarkers to Cimaglermin Alfa (GGF2) Clinical Trials

“…The absolute values for the increase in both M30 (cc‐K18) and M65 (cc‐K18 and FL‐K18) were higher than previously reported in healthy adult volunteers receiving recurrent therapeutic doses of acetaminophen12 but were significantly lower than those observed in patients with serious DILI due to acetaminophen overdose 15. The mathematical ratio of M30 to M65 is termed the “apoptotic index” and has been recently proposed to estimate the relative proportions of cell death by apoptosis and necrosis 4. Across all subjects with elevated cc‐K18 and FL‐K18, the apoptotic index (calculated using peak cc‐K18 and peak FL‐K18 values for each subject) ranged from 0.53–0.76, suggesting that apoptosis was the major form of cell death.…”

“…In contrast, during apoptosis, caspase cleavage of K18 occurs resulting in cc‐K18 release from apoptotic cells after loss of membrane integrity. The ratio of cc‐K18 to FL‐K18 (the “apoptotic index”) has been proposed to estimate the relative contributions of necrosis and apoptosis to cell death 4. Macrophage colony‐stimulating factor (CSF‐1) represents a proregenerative biomarker that has been shown to increase following partial hepatectomy in mice and humans and to correlate with survival in acetaminophen‐induced hepatotoxicity 5…”

The antiapoptotic effect of IL-6 autocrine loop in a cellular model of advanced prostate cancer is mediated by Mcl-1

“…CK18 is expressed and released by most types of carcinomas, including that of the prostate. In the case of apoptosis, soluble CK18 has been demonstrated to be a substrate for caspases (Kramer et al, 2004). As shown in Figure 1, exposure to the cytokine induced a 100 and 75% increase of apoptosis in parental and LNCaP-IL-6À cells, respectively, whereas no changes in cleaved CK18 levels were observed in IL-6-treated LNCaP-IL-6 þ cells.…”

Serum cytokeratin 18 as a biomarker for gastric cancer

“…In addition, many patients with advanced gastric cancer have normal levels of these markers. Classical biomarkers of gastric cancer do not have sufficient 15 sensitivity for population screening. In the present study, diagnostic sensitivities of M30 (67.5%) and M65 (70.1%) were higher than those of classical tumor markers (CEA and CA19-9, both 22.2%).…”

“…Enzyme-linked immunosorbent assays (ELISAs) have been developed to measure the circulating levels of caspase-cleaved and total soluble CK18 fragments. M30 and M65 antibodies can be used in ELISA to detect the different forms of CK18 released into circulation during caspase-dependent or total cell death, respectively, and these antibodies are potential biomarkers of epithelial cancers [15]. Activated caspases 3, 7, and 9 cleave CK18 during apoptosis, which leads to collapse of the cytoskeleton and subsequent formation of apoptotic bodies; subsequently, the proteolytic fragments of caspase-cleaved CK18 are released into circulation [16,17].…”