The association between diastolic blood pressure and massive transfusion in severe trauma: a retrospective single-center study.

OBJECTIVE: To evaluate the association between diastolic blood pressure and massive transfusion in severe trauma. Method: The retrospective study was conducted at a tertiary emergency medical centre in Gwangju , Republic of Korea, and comprised data of severe trauma patients with injury severity score >15 presenting between January 2016 and December 2017. Multivariate logistic regression analysis was performed to evaluate the association between diastolic blood pressure and massive transfusion. Receiver operating characteristic curve analysis was performed to estimate the prognostic performance of diastolic blood pressure. Data was analysed using SPSS 18. RESULTS: Of the 827 patients, 64(7.7%) underwent massive transfusion. After adjusting the confounders, diastolic blood pressure was found to be an independent factor in predicting massive transfusion (odds ratio: 0.965; 95% confidence interval: 0.956-0.975). CONCLUSIONS: Initially low diastolic blood pressure was found to be an independent predictor for massive transfusion in severe trauma cases.

Hepatic stellate cell-specific knockout of transcriptional intermediary factor 1γ aggravates liver fibrosis.

Transforming growth factor ß (TGFß) is a crucial factor in fibrosis, and transcriptional intermediary factor 1γ (TIF1γ) is a negative regulator of the TGFß pathway; however, its role in liver fibrosis is unknown. In this study, mesenchymal stem cells derived from human embryonic stem cells (hE-MSCs) that secrete hepatocyte growth factor (HGF) were used to observe the repair of thioacetamide (TAA)-induced liver fibrosis. Our results showed that TIF1γ was significantly decreased in LX2 cells when exposed to TGFß1. Such decrease of TIF1γ was significantly prevented by co-culture with hE-MSCs. Interaction of TIF1γ with SMAD2/3 and binding to the promoter of the α-smooth muscle gene (αSMA) suppressed αSMA expression. Phosphorylation of cAMP response element-binding protein (CREB) and binding on the TIF1γ promoter region induced TIF1γ expression. Furthermore, hepatic stellate cell-specific TIF1γ-knockout mice showed aggravation of liver fibrosis. In conclusion, loss of TIF1γ aggravates fibrosis, suggesting that a strategy to maintain TIF1γ during liver injury would be a promising therapeutic approach to prevent or reverse liver fibrosis.

Biallelic Deletion of Pxdn in Mice Leads to Anophthalmia and Severe Eye Malformation.

Peroxidasin (PXDN) is a unique peroxidase containing extracellular matrix motifs and stabilizes collagen IV networks by forming sulfilimine crosslinks. PXDN gene knockout in Caenorhabditis elegans (C. elegans) and Drosophila results in the demise at the embryonic and larval stages. PXDN mutations lead to severe eye disorders, including microphthalmia, cataract, glaucoma, and anterior segment dysgenesis in humans and mice. To investigate how PXDN loss of function affects organ development, we generated Pxdn knockout mice by deletion of exon 1 and its 5' upstream sequences of the Pxdn gene using the CRISPR/Cas9 system. Loss of both PXDN expression and collagen IV sulfilimine cross-links was detected only in the homozygous mice, which showed completely or almost closed eyelids with small eyes, having no apparent external morphological defects in other organs. In histological analysis of eye tissues, the homozygous mice had extreme defects in eye development, including no eyeballs or drastically disorganized eye structures, whereas the heterozygous mice showed normal eye structure. Visual function tests also revealed no obvious functional abnormalities in the eyes between heterozygous mice and wild-type mice. Thus, these results suggest that PXDN activity is essential in eye development, and also indicate that a single allele of Pxdn gene is sufficient for eye-structure formation and normal visual function.

Thymosin ß-4 is a novel regulator for primary cilium formation by nephronophthisis 3 in HeLa human cervical cancer cells.

Thymosinß-4(Tß4) is an actin-sequestering protein involved in tumor malignancy. Primary cilia, microtubule-based organelles, are present in most eukaryotic cells, which might be related to tumor cell transformation. Here, we investigated whether ciliogenesis is affected by Tß4 in HeLa human cervical cancer cells. The inhibition of Tß4 attenuated primary cilia formation. The frequency of cilia was increased by Tß4 overexpression. When yeast two-hybrid assay was performed by using Tß4 as a bait, we rescued nephronophthisis 3(NPHP3), one of the components of primary cilia. Interaction of Tß4 with NPHP3 in mammalian cells was confirmed by GST-pulldown assay. Their intracellular co-localization was observed by immunofluorescence staining at peripheral surface of cells. In addition, the number of ciliated cells was reduced by the inhibition of NPHP3. Moreover, NPHP3 expression was decreased by the inhibition of Tß4 but it was increased by Tß4 overexpression. Taken together, the results demonstrate that primary cilia formation could be regulated by Tß4 through its interaction with NPHP3 and/or the control of NPHP3 expression. It suggests that Tß4 is a novel regulator for primary cilia formation by NPHP3. It also suggests that tumorigenesis could be associated with inappropriate regulation of Tß4 and/or NPHP3 expression to maintain primary cilia formation normally.

Prognostic Performance Evaluation of the International Society on Thrombosis and Hemostasis and the Korean Society on Thrombosis and Hemostasis Scores in the Early Phase of Trauma.

BACKGROUND: Disseminated intravascular coagulation (DIC) contributes to poor outcome in the early phase of trauma. We aimed to analyze and compare the prognostic performances of the International Society on Thrombosis and Hemostasis (ISTH) and the Korean Society on Thrombosis and Hemostasis (KSTH) scores in the early phase of trauma. METHODS: Receiver operating characteristics analysis was used to examine the prognostic performance of both scores, and multivariate analysis was used to estimate the prognostic impact of the ISTH and KSTH scores in the early phase of trauma. The primary outcome was 24-hour mortality and the secondary outcome was massive transfusion. RESULTS: Of 1,229 patients included in the study, the 24-hour mortality rate was 7.6% (n = 93), and 8.1% (n = 99) of patients who received massive transfusions. The area under the curves (AUCs) of the KSTH and ISTH scores for 24-hour mortality were 0.784 (95% confidence interval [CI], 0.760-0.807) and 0.744 (95% CI, 0.718-0.768), respectively. The AUC of KSTH and ISTH scores for massive transfusion were 0.758 (95% CI, 0.734-0.782) and 0.646 (95% CI, 0.619-0.673), respectively. The AUCs of the KSTH score was significantly different from those of the ISTH score. Overt DIC according to KSTH criteria only, was independently associated with 24-hour mortality (odds ratio [OR], 2.630; 95% CI, 1.456-4.752). Only the KSTH score was independently associated with massive transfusion (OR, 1.563; 95% CI, 1.182-2.068). CONCLUSION: The KSTH score demonstrates a better prognostic performance for outcomes than the ISTH score in the early phase of trauma.

BIS-mediated STAT3 stabilization regulates glioblastoma stem cell-like phenotypes.

Glioblastoma stem cells (GSCs) are a subpopulation of highly tumorigenic and stem-like cells that are responsible for resistance to conventional therapy. Bcl-2-intreacting cell death suppressor (BIS; also known as BAG3) is an anti-apoptotic protein that is highly expressed in human cancers with various origins, including glioblastoma. In the present study, to investigate the role of BIS in GSC subpopulation, we examined the expression profile of BIS in A172 and U87-MG glioblastoma cell lines under specific in vitro culture conditions that enrich GSC-like cells in spheres. Both BIS mRNA and protein levels significantly increased under the sphere-forming condition as compared with standard culture conditions. BIS depletion resulted in notable decreases in sphere-forming activity and was accompanied with decreases in SOX-2 expression. The expression of STAT3, a master regulator of stemness, also decreased following BIS depletion concomitant with decreases in the nuclear levels of active phosphorylated STAT3, while ectopic STAT3 overexpression resulted in recovery of sphere-forming activity in BIS-knockdown glioblastoma cells. Additionally, immunoprecipitation and confocal microscopy revealed that BIS physically interacts with STAT3. Furthermore, BIS depletion increased STAT3 ubiquitination, suggesting that BIS is necessary for STAT3 stabilization in GSC-like cells. BIS depletion also affected epithelial-to-mesenchymal transition-related genes as evidenced by decrease in SNAIL and MMP-2 expression and increase in E-cadherin expression in GSC-like cells. Our findings suggest that high levels of BIS expression might confer stem-cell-like properties on cancer cells through STAT3 stabilization, indicating that BIS is a potential target in cancer therapy.

The synergistic effect of combination temozolomide and chloroquine treatment is dependent on autophagy formation and p53 status in glioma cells.

Temozolomide (TMZ) is an alkylating agent used for the treatment of glioblastoma. The late autophagy inhibitor chloroquine (CQ) inhibits glioblastoma tumors in a p53-independent and p53-dependent manner. We addressed a possible beneficial effect of combination treatment with TMZ and CQ by examining the molecular and cellular mechanism of co-treatment. Combination treatment of U87 cell (wild type p53) with TMZ and CQ synergistically reduced cell proliferation and enhanced apoptosis, with increased sub-G1 hypodiploid cells and caspase activation. This effect was abolished by a pan-caspase inhibitor, Z-VAD-FMK. TMZ induced autophagy, and the addition of CQ further increased autophagic vacuoles. Inhibition of early stages of autophagy by Beclin 1 knockdown and 3-methyladenine pretreatment prevented the enhanced effect of the combination treatment. The combination treatment also upregulated p53 and phospho-p53 levels, whereas p53 knockdown or overexpression of mutant p53 abolished the combination effect. In contrast, combination therapy had no enhanced effect on U373 cell (mutant p53) proliferation and apoptosis within 3 d, although TMZ induced autophagy and co-treatment with CQ increased autophagic vacuole accumulation. However, long term combination treatment for 9-10 d effectively decreased clonal and cellular growth with increased G2-M arrest. This effect was also abolished by Beclin 1 knockdown. Our data support the beneficial effect of combination treatment with TMZ and CQ in glioma via differential autophagy-associated mechanisms, depending on p53 status.

Inositol Polyphosphate-5-Phosphatase F (INPP5F) inhibits STAT3 activity and suppresses gliomas tumorigenicity.

Glioblastoma (GBM), the most common type of primary malignant brain tumors harboring a subpopulation of stem-like cells (GSCs), is a fast-growing and often fatal tumor. Signal Transducer and Activator of Transcription 3 (STAT3) is one of the major signaling pathways in GSCs maintenance but the molecular mechanisms underlying STAT3 deregulation in GSCs are poorly defined. Here, we demonstrate that Inositol Polyphosphate-5-Phosphatase F (INPP5F), one of the polyphosphoinositide phosphatases, is differentially expressed in GSCs from glioma patients, and is identified as an inhibitor of STAT3 signaling via interaction with STAT3 and inhibition of its phosphorylation. Constitutively expressed INPP5F showed to suppress self-renewal and proliferation potentials of glioblastoma cells and reduced tumorigenicity of glioblastoma. In addition, loss of INPP5F gene in gliomas is significantly correlated with lower overall patient survivals. These findings suggest that INPP5F is a potential tumor suppressor in gliomas via inhibition of STAT3 pathway, and that deregulation of INPP5F may lead to contribution to gliomagenesis.

Identification of molecular pathways facilitating glioma cell invasion in situ.

Gliomas are mostly incurable secondary to their diffuse infiltrative nature. Thus, specific therapeutic targeting of invasive glioma cells is an attractive concept. As cells exit the tumor mass and infiltrate brain parenchyma, they closely interact with a changing micro-environmental landscape that sustains tumor cell invasion. In this study, we used a unique microarray profiling approach on a human glioma stem cell (GSC) xenograft model to explore gene expression changes in situ in Invading Glioma Cells (IGCs) compared to tumor core, as well as changes in host cells residing within the infiltrated microenvironment relative to the unaffected cortex. IGCs were found to have reduced expression of genes within the extracellular matrix compartment, and genes involved in cell adhesion, cell polarity and epithelial to mesenchymal transition (EMT) processes. The infiltrated microenvironment showed activation of wound repair and tissue remodeling networks. We confirmed by protein analysis the downregulation of EMT and polarity related genes such as CD44 and PARD3 in IGCs, and EFNB3, a tissue-remodeling agent enriched at the infiltrated microenvironment. OLIG2, a proliferation regulator and glioma progenitor cell marker upregulated in IGCs was found to function in enhancing migration and stemness of GSCs. Overall, our results unveiled a more comprehensive picture of the complex and dynamic cell autonomous and tumor-host interactive pathways of glioma invasion than has been previously demonstrated. This suggests targeting of multiple pathways at the junction of invading tumor and microenvironment as a viable option for glioma therapy.

Myocilin is involved in NgR1/Lingo-1-mediated oligodendrocyte differentiation and myelination of the optic nerve.

Myocilin is a secreted glycoprotein that belongs to a family of olfactomedin domain-containing proteins. Although myocilin is detected in several ocular and nonocular tissues, the only reported human pathology related to mutations in the MYOCILIN gene is primary open-angle glaucoma. Functions of myocilin are poorly understood. Here we demonstrate that myocilin is a mediator of oligodendrocyte differentiation and is involved in the myelination of the optic nerve in mice. Myocilin is expressed and secreted by optic nerve astrocytes. Differentiation of optic nerve oligodendrocytes is delayed in Myocilin-null mice. Optic nerves of Myocilin-null mice contain reduced levels of several myelin-associated proteins including myelin basic protein, myelin proteolipid protein, and 2'3'-cyclic nucleotide 3'-phosphodiesterase compared with those of wild-type littermates. This leads to reduced myelin sheath thickness of optic nerve axons in Myocilin-null mice compared with wild-type littermates, and this difference is more pronounced at early postnatal stages compared with adult mice. Myocilin also affects differentiation of oligodendrocyte precursors in vitro. Its addition to primary cultures of differentiating oligodendrocyte precursors increases levels of tested markers of oligodendrocyte differentiation and stimulates elongation of oligodendrocyte processes. Myocilin stimulation of oligodendrocyte differentiation occurs through the NgR1/Lingo-1 receptor complex. Myocilin physically interacts with Lingo-1 and may be considered as a Lingo-1 ligand. Myocilin-induced elongation of oligodendrocyte processes may be mediated by activation of FYN and suppression of RhoA GTPase.

Histone demethylase Jumonji D3 (JMJD3) as a tumor suppressor by regulating p53 protein nuclear stabilization.

Histone methylation regulates normal stem cell fate decisions through a coordinated interplay between histone methyltransferases and demethylases at lineage specific genes. Malignant transformation is associated with aberrant accumulation of repressive histone modifications, such as polycomb mediated histone 3 lysine 27 (H3K27me3) resulting in a histone methylation mediated block to differentiation. The relevance, however, of histone demethylases in cancer remains less clear. We report that JMJD3, a H3K27me3 demethylase, is induced during differentiation of glioblastoma stem cells (GSCs), where it promotes a differentiation-like phenotype via chromatin dependent (INK4A/ARF locus activation) and chromatin independent (nuclear p53 protein stabilization) mechanisms. Our findings indicate that deregulation of JMJD3 may contribute to gliomagenesis via inhibition of the p53 pathway resulting in a block to terminal differentiation.

Gliomagenesis arising from Pten- and Ink4a/Arf-deficient neural progenitor cells is mediated by the p53-Fbxw7/Cdc4 pathway, which controls c-Myc.

Glioblastoma multiforme is the most common type of primary malignant brain tumor and may arise from a cell with neural stem-like properties. Deregulation of the retinoblastoma, phosphoinositide-3 kinase (PI3K), and p53 pathways are molecular hallmarks of this disease. Recent work has shown that p53(-/-)Pten(-/-) mice form gliomas in a c-Myc-dependent manner. To explore the role of the INK4A/ARF locus and Pten deletions in gliomagenesis, we generated Pten(-/-)Ink4a/Arf(-/-) mouse neural stem cells (mNSC) and such cells were highly proliferative, self-renewing, relatively refractory to differentiation, and induced both low- and high-grade glioma formation in vivo. In contrast to p53(-/-) Pten(-/-) mNSCs, however, Pten(-/-)Ink4a/Arf(-/-) mNSCs do not express appreciable levels of c-Myc in vitro, although glioma stem cells derived from thesecells did. Sequencing of Pten(-/-)Ink4a/Arf(-/-) mNSC-derived tumors revealed spontaneous mutations in Tp53 in vivo with subsequent downregulation of Fbxw7. Expression of p53 mutants in Pten(-/-)Ink4a/Arf(-/-) mNSC or knockdown of Fbxw7 resulted in reexpression of c-Myc with enhanced Pten(-/-)Ink4a/Arf(-/-) mNSC tumorigenecity. We propose that p53 mutations contribute to gliomagenesis by both allowing the overexpression of c-Myc through downregulation of Fbxw7 and by protecting against c-Myc-induced apoptosis.

Qualitative network modeling of the Myc-p53 control system of cell proliferation and differentiation.

A kinetic model of a molecular control system for the cellular decision to proliferate or differentiate is formulated and analyzed for the purpose of understanding how the system can break down in cancer cells. The proposed core of this control system is composed of the transcription factors Myc and p53. The network of interactions between these factors involves negative and positive feedback loops that are linked to pathways involved in differentiation, cell cycle, and apoptosis. Understanding the dynamics of the Myc-p53 control system is aided by the postulate that there exists a cancer zone defined as a range of oncogenic Myc activities where the probability of initiating cancer is high. We propose that an essential role of p53 is to prevent the system from entering or staying too long in the cancer zone by downregulating Myc or, when Myc activity somehow becomes too high, by inducing apoptosis, cell cycle arrest, or differentiation. Kinetic modeling illustrates how deletions or aberrations in PTEN, MDM2, and ARF (genes implicated in various cancers, including glioma) affect the Myc-p53 control system. In addition, computer simulations demonstrate how this control system generates different cellular phenotypes characterized by rates of cellular differentiation and proliferation.

A HIF-1 target, ATIA, protects cells from apoptosis by modulating the mitochondrial thioredoxin, TRX2.

The regulation of apoptosis is critical for controlling tissue homeostasis and preventing tumor formation and growth. Reactive oxygen species (ROS) generation plays a key role in such regulation. Here, we describe a HIF-1 target, Vasn/ATIA (anti-TNFα-induced apoptosis), which protects cells against TNFα- and hypoxia-induced apoptosis. Through the generation of ATIA knockout mice, we show that ATIA protects cells from apoptosis through regulating the function of the mitochondrial antioxidant, thioredoxin-2, and ROS generation. ATIA is highly expressed in human glioblastoma, and ATIA knockdown in glioblastoma cells renders them sensitive to hypoxia-induced apoptosis. Therefore, ATIA is not only a HIF-1 target that regulates mitochondrial redox pathways but also a potentially diagnostic marker and therapeutic target in human glioblastoma.

Mitochondrial microsatellite instability of colorectal cancer stroma.

Mitochondrial microsatellite instability (mtMSI) and mutations of mitochondrial DNA has been reported in cancer epithelia of carcinomas. However, mtMSI in cancer stroma has not yet been identified in human cancers. In this study, we attempted to determine if mtMSI occurs in the cancer stroma of sporadic colorectal cancers, and if the stromal mtMSI has any correlations with stromal nuclear MSI (nMSI) and cancer epithelial mtMSI. Nine microsatellite sequences within the D-loop and 5 coding genes for mtMSI, and 9 microsatellites for nMSI were analyzed in the microdissected cancer epithelia and adjacent stromas of 48 sporadic colorectal cancers. Overall, 23 somatic mitochondrial DNA alterations were detected in 15 cancer epithelia (31.2%) and 5 stromas (10.4%). The mutations consisted of 19 D-loop mtMSI alterations, and 1 missense and 3 framshift mutations of repeat sequences within the coding genes. All of the 5 stromal genetic alterations showed D-loop mtMSI. In regards to other MSI status, the stromal mtMSI had no association with stromal nMSI or epithelial mtMSI, either. These findings indicate that in addition to the cancer epithelia the cancer stroma harbor mtMSI, and suggest a possible role of stromal mtMSI in the pathogenesis of colorectal cancers. Furthermore, the data suggest that stromal mtMSI may occur independently of stromal nMSI and epithelial mtMSI in sporadic colorectal cancers.

Inactivating mutation of the pro-apoptotic gene BID in gastric cancer.

There is evidence that deregulation of apoptosis is mechanistically involved in cancer development and somatic mutations of apoptosis-related genes have been reported in human cancers. BID, a pro-apoptotic member of the Bcl-2 family, interconnects the extrinsic apoptosis pathway initiated by death receptors to the intrinsic apoptosis pathway. To explore the possibility that genetic alterations of BID might be involved in the development of human cancers, this study analysed the entire coding region and all splice sites in the human BID gene in 67 advanced gastric carcinomas. Overall, four BID mutations (6.0%) were detected that consisted of one frameshift and three missense mutations. The tumour-derived BID mutants were expressed in 293T cells and it was found that, compared with wild-type BID, the frequency of apoptosis was significantly reduced in cells expressing the gene containing the frameshift mutation. Furthermore, expression of the inactivating frameshift mutant interfered with cell death by overexpression of death receptors, indicating that this mutant inhibits the extrinsic apoptosis pathway in a dominant-negative fashion. Also, the frameshift mutation rendered cancer cells resistant to apoptosis induced by the anti-cancer drug 5-fluorouracil (5-FU). This is the first report of BID gene mutation in human malignancy. The data suggest that such mutations occur rarely in gastric cancers and that only a small fraction of BID mutations may lead to the loss of its apoptotic function.

Inactivating mutations of proapoptotic Bad gene in human colon cancers.

Evidence exists that deregulation of apoptosis is involved in the mechanisms of cancer development, and the somatic mutations of apoptosis-related genes have been reported in human cancers. Bcl-XL/Bcl-2-associated death promoter (Bad), a proapoptotic member of Bcl-2 family, plays an important role in the intrinsic apoptosis pathway. To explore the possibility that the genetic alterations of Bad might be involved in the development of human cancers, we analyzed the entire coding region and all splice sites of human Bad gene in 47 colon adenocarcinomas. Overall, we detected two somatic missense mutations (4.3%) in Bad gene. Interestingly, both of the Bad mutations were detected in the gene sequences encoding the Bcl-2 homology3 domain of Bad, which has a crucial role in inducing cell death. Transfection study revealed that both of the tumor-derived Bad mutants had decreased apoptosis activities compared with the wild-type Bad, indicating that the Bad mutations reduced the cell death function of Bad. Co-immunoprecipitation assay revealed that binding of one of the tumor-derived Bad mutants with Bcl-2 and Bcl-XL is reduced. This is the first report on Bad gene mutation in human malignancies, and our data suggest that Bad gene is occasionally mutated in colon cancers and that somatic mutation of Bad may contribute to the development of colon cancers.

BRAF and KRAS mutations in stomach cancer.

Ras proteins control signaling pathways that are key regulators of several aspects of normal cell growth and malignant transformation. BRAF, which encodes a RAF family member in the downstream pathway of RAS, is somatically mutated in a number of human cancers. The activating mutation of BRAF is known to play a role in tumor development. As there have been no data on the BRAF mutation in stomach cancer, we analysed the genomic DNAs from 319 stomach carcinomas for the detection of somatic mutations of BRAF. Overall, we detected BRAF mutations in seven stomach carcinomas (2.2%). Five of the seven BRAF mutations involved Val 599, the previously identified hotspot, but the substituted amino acid (V599 M) was different from the most common BRAF mutation (V599E). The remaining two mutations involved a conserved amino acid (D593G). One tumor had both BRAF and KRAS mutations. This is the first report on BRAF mutation in stomach cancer, and the data indicate that BRAF is occasionally mutated in stomach cancer, and suggest that alterations of RAS pathway both by RAS and BRAF mutations contribute to the pathogenesis of stomach cancer.

Mutational analysis of Noxa gene in human cancers.

There has been mounting evidence that dysregulation of apoptosis is involved in the mechanisms of cancer development and somatic mutations of apoptosis-related genes have been reported in human cancers. Noxa, a Bcl-2 homology 3 (BH3)-only member of the Bcl-2 family, is known to interact with anti-apoptotic Bcl-2 family members and induces apoptosis. The aim of this study was to explore the possibility that the Noxa gene is mutated in human cancers. We have analyzed the entire coding region and all splice sites of the Noxa gene for the detection of somatic mutations in a series of human cancers, including carcinomas from stomach, colon, liver, urinary bladder and lung by polymerase chain reaction (PCR), single strand conformation polymorphism (SSCP), and DNA sequencing. We found one somatic mutation of the Noxa gene in a transitional cell carcinoma (TCC) of the urinary bladder. To evaluate the functional alterations of the mutant in apoptosis, we overexpressed the mutant and wild-type Noxa in 293T and HeLa cells, but could not find any significant difference in cell death between the wild-type and mutant Noxa. These data suggest that Noxa is rarely mutated in human carcinomas and that the contribution of Noxa gene mutation in the pathogenesis of human cancer might not be related to cell death mechanisms.

Inactivating mutations of caspase-8 gene in colorectal carcinomas.

BACKGROUND & AIMS: There has been evidence that dysregulation of apoptosis is involved in the pathogenesis of cancer development. Caspase-8 is an initiation caspase that activates the caspase cascade during apoptosis. The aim of this study was to explore the possibility that mutation of the caspase-8 gene might be involved in the development of colorectal cancer. METHODS: We analyzed the entire coding region of the caspase-8 gene for the detection of somatic mutations in 180 colorectal tumors (98 invasive carcinomas and 82 adenomas) by polymerase chain reaction, single-strand conformation polymorphism, and DNA sequencing. RESULTS: Overall, we detected a total of 5 somatic mutations in 98 invasive carcinomas (5.1%), but no mutations were detected in 82 adenomas (0%). The frequency of caspase-8 mutation in the carcinomas was significantly higher than that in adenomas (P < 0.05). The 5 mutations consisted of 1 frameshift, 1 nonsense mutation, and 3 missense mutations. We expressed the 5 tumor-derived caspase-8 mutants and found that 3 of the 5 mutations markedly decreased apoptosis activity of caspase-8. Furthermore, expression of the inactivating caspase-8 mutants interfered with apoptosis by death receptor overexpression, indicating that these mutants have dominant-negative inhibition of the death receptor-induced apoptosis. CONCLUSIONS: The presence of caspase-8 mutation in colon carcinomas suggests that caspase-8 gene mutation might lead to the loss of its apoptotic function and contribute to the pathogenesis of colorectal carcinomas, especially at the late stage of colorectal carcinogenesis.