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Protein kinase C (PKC) is a novel member of the PKC family that differs from the other isozymes in structural and biochemical properties. The precise function of PKC is not known. The present studies demonstrate that PKC is cleaved during apoptosis induced by 1-␤-D-arabinofuranosylcytosine (ara-C) and other genotoxic agents. PKC cleavage is blocked in cells that overexpress the anti-apoptotic Bcl-x L protein or the baculovirus p35 protein. Our results demonstrate that PKC is cleaved by caspase-3 at the CQND 378 S site. Cleavage of PKC is associated with release of the catalytic domain and activation of its kinase function. We also show that, unlike the cleaved fragments of PKC␦ and , overexpression of the PKC catalytic domain is not lethal. Cells stably expressing the catalytic fragment of PKC, however, are more sensitive to apoptosis induced by genotoxic stress. In addition, expression of the caspase-resistant PKC mutant partially inhibits DNA damageinduced apoptosis. These findings demonstrate that PKC is cleaved by caspase-3 and that expression of the catalytic domain sensitizes cells to the cytotoxic effects of ara-C and other anticancer agents.

Metastasis-associated protein MTA1 and histone deacetylase form a protein complex with histone deacetylase activity that plays an important role in histone deacetylation, alteration of chromatin structure and transcriptional control. The precise role of the MTA1 protein in the malignant progression of human cancers remains unknown, however, especially its overexpression and relationship with histone acetylation/ deacetylation in experimental and clinical tumors. The expression levels of MTA1 protein and the acetylation levels of histone H4 were examined in 70 cases of surgically resected esophageal squamous cell carcinomas, using immunohistochemistry. The intensities of immunostaining of MTA1 protein and acetylated histone H4 in carcinoma tissues (Ca) were compared to normal epithelium (N) contained in the same section. Thirty of 70 cases (42.9%) displayed overexpression of MTA1 protein (N < Ca). Cancers overexpressing MTA1 protein invaded deeper into the esophageal wall (p < 0.005) and showed significantly higher degrees of lymph node metastasis (p < 0.01), higher pathological stage, more lymphatic involvement and poorer prognosis (p < 0.05) than the remaining cases. The acetylation levels of histone H4 inversely correlated to the depth of cancer invasion and pathological stage (p < 0.05), and the patients with higher level of histone H4 acetylation had a better prognosis (p < 0.05). Furthermore, immunostaining patterns of MTA1 and acetylated histone H4 were inversely correlated (p < 0.001), demonstrating the relationship of deacetylation of histone H4 in MTA1-overexpressing carcinomas. In conclusion, the data suggest that the overexpression of MTA1 protein and acetylation level of histone H4 protein, both of which are closely related, might be useful predictors of malignant potential of esophageal squamous cell carcinomas. Thus, strategies involving inhibition of MTA1 function as well as inhibition of histone deacetylation could be novel approaches for the treatment of esophageal squamous cell carcinomas. © 2004 Wiley-Liss, Inc. Key words: esophageal squamous cell carcinoma; metastasis-associated protein MTA1; histone deacetylation; cancer progression; prognosisModifications of chromatin structure by acetylation/deacetylation of the histone proteins play a central role in the regulation of transcription. 1,2 The levels of acetylation/deacetylation of histone proteins are determined by 2 opposing enzymes, histone acetyltransferases (HATs) and histone deacetylases (HDACs), especially HDAC1 and HDAC2. 2 These alterations of chromatin structure by HATs and HDACs have been implicated in the process of carcinogenesis. 3,4 For example, the retinoblastoma protein pRB has been shown to have the ability to associate with HDAC1 and recruit the pRB complex to the E2F-regulated promoters of genes such as cell-cycle regulatory genes, resulting in inhibition of E2-regulated gene expression. Thus, the pRB/HDAC1 complex is likely to be a key element in the control of cell proliferation and differentiation. 4 -6 HDAC inhibito...

The IAP (inhibitor of apoptosis) family of anti-apoptotic proteins regulates programmed cell death. Of the six known human IAP-related proteins, XIAP is the most potent inhibitor. To study the mechanistic effects of XIAP on DNA damage-induced apoptosis, we prepared U-937 cells that stably overexpress XIAP. The results demonstrate that XIAP inhibits apoptosis induced by 1-[␤-D-arabinofuranosyl]cytosine (ara-C) and other genotoxic agents. XIAP had no detectable effect on ara-C-induced release of mitochondrial cytochrome c and attenuated cleavage of procaspase-9. In addition, we show that ara-C induces the association of XIAP with the cleaved fragments of caspase-9 and thereby inhibition of caspase-9 activity. The results also demonstrate that ara-C induces cleavage of procaspase-3 by a caspase-8-dependent mechanism and that XIAP inhibits caspase-3 activity. These results demonstrate that XIAP functions downstream of procaspase-9 cleavage as an inhibitor of both proteolytically processed caspase-9 and -3 in the cellular response to genotoxic stress.

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Objective: An increase in the activity of mitogen-activated protein kinases (MAPKs) has been correlated with a more malignant phenotype in several tumor models in vivo. This study was designed to clarify the expression of MKP-1 in surgically resected hepatocellular carcinoma (HCC). Methods: We reviewed the cases of 77 patients who had undergone initial liver resection for HCC without preoperative treatment. Immunohistochemical analysis of MKP-1 was performed on paraffin-embedded tissues. The correlation between MKP-1 expression and clinical outcome was investigated. Results: Tumor cells were immunohistochemically stained for MKP-1 expression, and the same levels as in normal hepatocytes were detected in 66 (85%) of 77 HCC patients, being decreased in 11 (15%) HCCs. Decreased MKP-1 expression significantly correlated with serum α-fetoprotein levels and tumor size (p < 0.05). The disease-free survival rates in MKP-1-negative and -positive patients were 0 and 31.0% at 5 years, respectively (p < 0.01). The survival rates after a surgical resection in MKP-1-negative and -positive patients were 18.2 and 65.5% at 5 years, respectively (p < 0.01). Conclusions: The MKP-1 expression in HCC was an independent prognostic factor for outcome in HCC patients. In the future, it will be useful to explore whether the phosphatase expression might account for the response to HCC treatments targeting at MAPK activation.

Abstract. Second mitochondria-derived activator of caspases/ direct inhibitor of apoptosis-binding protein with low pI (Smac/DIABLO) is released by mitochondria in response to apoptotic stimuli and is thought to regulate apoptosis by antagonizing inhibitors of apoptosis proteins, which play an important role in sensitization of cancer cells to various therapeutic regimens. The expression of Smac/DIABLO has been demonstrated in various cancer cells, though little is known about its clinical significance with respect to colorectal cancer. The current study was designed to evaluate the relationship between prognosis and Smac/DIABLO expression by clinicopathological analysis of patients with colorectal cancer. Smac/DIABLO expression was evaluated using immunohistochemical staining in 121 consecutive patients with colorectal cancer and the relationship between Smac/DIABLO expression and clinicopathological factors was analyzed. Smac/DIABLO-positive expression was detected in 80 of the 121 patients (66%). The incidence of lymph node and distant metastasis in Smac/DIABLOnegative cancer was significantly higher than that in Smac/DIABLO-positive cancer (P=0.0004 and P=0.003, respectively). While univariate analysis showed that survival in patients with Smac/DIABLO-negative expression was significantly poorer than in Smac/DIABLO-positive cases (P<0.0001), Smac/DIABLO-negative expression was a prognostic indicator independent of Dukes' staging and lymph node metastasis by multivariate analysis. This study proposes that the decrease of Smac/DIABLO expression is an independent factor determining the poorer prognosis of patients with colorectal cancer.