The role of caspase 3 in producing cytokeratin 19 fragment (CYFRA21-1) in human lung cancer cell lines

Abstract

The CYFRA 21‐1 assay, which detects cytokeratin 19 (CK19) fragment, is widely used as a tumor marker for lung cancer, particularly non‐small cell lung cancer. However, the reason that some lung cancer cell lines release CYFRA 21‐1 in culture supernatants and others do not remains unclear. We hypothesized that the release of CYFRA 21‐1 might be related to the expression of CK19 and caspase 3. In order to prove this, the quantities of mRNA for CK19 were evaluated by the competitive reverse transcriptase‐polymerase chain reaction (RT‐PCR). CK19 protein synthesis was also evaluated by Western blotting and immunohistochemistry, and the levels of CYFRA 21‐1 in the culture supernatant were measured by an immunoradiometric assay. The expression of mRNA for caspase 3 was evaluated by the RT‐PCR, and caspase 3 protein synthesis was also evaluated by immunohistochemistry. In 13 lung cancer cell lines, the amounts of mRNA for CK19 correlated with the levels of CYFRA 21‐1 in culture supernatants, results of Western blotting for CK19, and positivities of immunohistochemistry for CK19. In 5 cell lines that produced a significant amount of CYFRA 21‐1, the level of CYFRA 21‐1 correlated with the positivity of RT‐PCR for caspase 3 and immunohistochmistry for caspase 3. This suggests that caspase 3 played a role in the formation of CYFRA 21‐1. In addition, the specific inhibitor of caspase 3 significantly inhibited the release of CYFRA 21‐1 in culture supernatants. In conclusion, we demonstrate that caspase 3, which cleaves several intermediate filaments and carries out cell apoptosis, played an important role in producing CYFRA 21‐1 in human lung cancer cell lines. © 2001 Wiley‐Liss, Inc.

Cytokeratin 8: The Dominant Type II Intermediate Filament Protein in Lung Cancer

“…Using MCF-7 cells, Sheard et al revealed TPS (i.e., CK18) and Cyfra 21-1 (CK19) are abundantly released during apoptosis and the release precedes DNA fragmentation (Sheard et al, 2002). We also observed mRNA expression of CK19 is essential in Cyfra 21-1 formation and caspase 3 plays an important role in producing Cyfra 21-1 in human lung cancer cell lines (Dohmoto et al, 2000;Dohmoto et al, 2001). Kramer described that large amounts of CK18 are released into extracellular space during necrosis induced by oligomycin, and that release of CK18 is a marker of epithelial cell death and not a marker of apoptosis (Kramer et al, 2004).…”

“…Furthermore, a specific inhibitor of caspase 3 efficiently inhibited the release of Cyfra 21-1 into culture supernatants. Based on these observations we concluded that caspase 3 plays an important role in the production of Cyfra 21-1 in lung cancer cell lines (Dohmoto et al, 2001). Pujol et al evaluated Cyfra 21-1 and TPS in 405 lung cancer patients (91 SCLC and 314 NSCLC) and in 59 patients with non-malignant lung diseases.…”

“…CKs have been found in extracellular spaces both in vitro and in vivo, in conditioned culture medium to circulating human blood. CK release is observed during cell growth/cell divisions, after surgical intervention, and upon cell death induction (e.g., radiation therapy and/or cytotoxic chemotherapy) (Bauer et al, 1986;Bjorklund & Bjorklund, 1983;Brabon et al, 1984;Caulin et al, 1997;Dohmoto et al, 2000;Dohmoto et al, 2001;Sheard et al, 2002;Tinnemans et al, 1995).…”

Relevance of circulating tumor cells, extracellular nucleic acids, and exosomes in breast cancer

“…They are classified into two types: acidic type I (CK9-CK20) and basic type II (CK1-CK8) [99]. Epithelial cells are targets of caspase-mediated proteolysis during apoptosis [100]. CK19 is cleaved by caspase 3 and soluble fragments (CYFRA 21-1), released into extracellular space, have been detected in sera of cancer patients [62,101] and reportedly do not correlate with that of full-length CK19 mRNA [62].…”

Comparative analysis of proteins in the culture supernatants of human intestinal epithelial cells infected with the wild-type and rtxE mutant of Vibrio vulnificus

“…The 14-3-3 protein is externalized to the medium by epithelial cell types, although the function of this protein in V. vulnificus infectious disease remains to be clarified. PRX II belongs to a ubiquitous PRX family, which has been associated with multiple functions, including enhancing natural killer cell activity (29), increasing cell resistance to oxidative stress (30), protecting erythrocytes against oxidative stress (31), and anti-HIV activity (32). However, the function of PRX II in the context of V. vulnificus infectious disease also requires clarification.…”