Jung Young Lee

PIK3CA gene is frequently mutated in breast carcinomas and hepatocellular carcinomas

A recent report revealed that phosphoinositide-3-kinase, catalytic, alpha (PIK3CA) gene is somatically mutated in several types of human cancer, suggesting the mutated PIK3CA gene as an oncogene in human cancers. However, because the previous report focused the mutational search primarily on colon cancers, the data on PIK3CA mutations in other types of human cancers have been largely unknown. Here, we performed mutational analysis of the PIK3CA gene by polymerase chain reaction-single-strand conformation polymorphism assay in 668 cases of common human cancers, including hepatocellular carcinomas, acute leukemias, gastric carcinomas, breast carcinomas, and non-small-cell lung cancers. We detected PIK3CA somatic mutations in 26 of 73 hepatocellular carcinomas (35.6%), 25 of 93 breast carcinomas (26.9%), 12 of 185 gastric carcinomas (6.5%), one of 88 acute leukemias (1.1%), and three of 229 non-small-cell lung cancers (1.3%). Some of the PIK3CA mutations were detected in the early lesions of breast cancer carcinoma, hepatocellular carcinoma, and gastric carcinomas, suggesting that PIK3CA mutation may occur independent of stage of the tumors. The high incidence and wide distribution of PIK3CA gene mutation in the common human cancers suggest that alterations of lipid kinase pathway by PIK3CA mutations contribute to the development of human cancers.

Alterations of Fas (Apo-1/CD95) gene in non-small cell lung cancer.

Fas (Apo-1/CD95) is a cell-surface receptor involved in cell death signaling. The key role of the Fas system in negative growth regulation has been studied mostly within the immune system, and somatic mutations of Fas gene in cancer patients have been described solely in lymphoid-lineage malignancies. However, many non-lymphoid tumor cells have been found to be resistant to Fas-mediated apoptosis, which suggests that Fas mutations, one of the possible mechanisms for Fas-resistance, may be involved in the pathogenesis of non-lymphoid malignancies as well. In this study, we have analysed the entire coding region and all splice sites of the Fas gene for the detection of the gene mutations in 65 human non-small cell lung cancers by polymerase chain reaction, single strand conformation polymorphism and DNA sequencing. Overall, five tumors (7.7%) were found to have the Fas mutations, which were all missense mutations. Four of the five mutations identified were located in the cytoplasmic region (death domain) known to be involved in the transduction of an apoptotic signal and one mutation was located in the transmembrane domain. This is the first report on the Fas gene mutations in non-lymphoid malignancies, and the data presented here suggests that alterations of the Fas gene might lead to the loss of its apoptotic function and contribute to the pathogenesis of some human lung cancers.

Increased expression of histone deacetylase 2 is found in human gastric cancer.

Accumulated evidence has established that aberrant regulation of histone deacetylases (HDACs) is one of the major causes of the development of human malignancies. Among different iso‐enzymes of HDAC and sirtuins grouped as the HDAC super family, little is known as to how histone deacetylase 2 (HDAC2) causes carcinogenesis in solid tumors. Here, in order to investigate the possible role of HDAC2 in gastric carcinogenesis, we analyzed the expression of HDAC2 in 71 gastric adenocarcinomas by immunohistochemistry. Moderate to strong expression of HDAC2 was found in 44 (62%) out of a total of 71 tumors. The majority of positive tumors, which were detected in the nucleus but not in normal gastric epithelium, did not express HDAC2 or showed only weak positive staining. Interestingly, we also noted that HDAC2 expression appeared to be associated with tumor aggressiveness as HDAC2 expression was observed to be statistically significant in advanced gastric cancer (P=0.0023, Chi‐square test) and in positive lymph node metastasis (P=0.0713, Chi‐square test). Taken together, these results suggest that HDAC2 may play an important role in the aggressiveness of gastric cancer.

Somatic Mutations of EGFR Gene in Squamous Cell Carcinoma of the Head and Neck

Purpose: Recently, the kinase domain mutations of epidermal growth factor receptor (EGFR) gene have been identified in non–small-cell lung cancer, and these mutations have been related to the clinical response to the tyrosine kinase inhibitor gefitinib. Gefitinib treatment has also shown clinical benefits in squamous cell carcinoma of the head and neck (SCCHN). The aim of this study was to explore the possibility that SCCHN harbored the EGFR mutations. Experimental Design: In this study, we analyzed EGFR gene in 41 SCCHN for the detection of the somatic mutations by PCR-single-strand conformational polymorphism analysis. Results: Overall, we detected three EGFR mutations (7.3%), and all of the mutations were the same in-frame deletion mutation in exon 19 (E746_A750del). Conclusion: These data indicated that in addition to non–small-cell lung cancer, SCCHN harbors the EGFR gene mutations, and suggested the rationale for the clinical applicability of gefinitib to SCCHN patients.

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.

Somatic Mutations of ERBB2 Kinase Domain in Gastric, Colorectal, and Breast Carcinomas

Purpose: Recent reports revealed that the kinase domain of the ERBB2 gene is somatically mutated in lung adenocarcinoma, suggesting the mutated ERBB2 gene as an oncogene in human cancers. However, because previous reports focused the mutational search of ERBB2 primarily on lung cancers, the data on ERBB2 mutations in other types of human cancers have been largely unknown. Experimental Design: Here, we did a mutational analysis of the ERBB2 kinase domain by PCR single-strand conformational polymorphism assay in gastric, colorectal, and breast carcinoma tissues. Results: We detected the ERBB2 kinase domain mutations in 9 of 180 gastric carcinomas (5.0%), in 3 of 104 colorectal carcinomas (2.9%), and in 4 of 94 breast carcinomas (4.3%). All of the detected ERBB2 mutations except for one in-frame deletion mutation were missense mutations. Of the 16 ERBB2 mutations detected, 4 affected Val777 in the exon 20 site, and 3 affected Leu755 in the exon 19 site. We simultaneously analyzed the somatic mutations of EGFR, K-RAS, PIK3CA, and BRAF genes in the 16 samples with ERBB2 mutations, and found that all of the 3 colorectal carcinoma samples with ERBB2 mutations harbored K-RAS mutations. Conclusion: This study showed that in addition to lung adenocarcinomas, ERBB2 kinase domain mutation occurs in other common human cancers such as gastric, breast, and colorectal cancers, and suggested that alterations of ERBB2-mediated signaling pathway by ERBB2 mutations alone or together with K-RAS mutations may contribute to the development of human cancers.

Sirtuin7 oncogenic potential in human hepatocellular carcinoma and its regulation by the tumor suppressors MiR-125a-5p and MiR-125b.

Sirtuins are nicotinamide adenine dinucleotide oxidized form (NAD+)‐dependent deacetylases and function in cellular metabolism, stress resistance, and aging. For sirtuin7 (SIRT7), a role in ribosomal gene transcription is proposed, but its function in cancer has been unclear. In this study we show that SIRT7 expression was up‐regulated in a large cohort of human hepatocellular carcinoma (HCC) patients. SIRT7 knockdown influenced the cell cycle and caused a significant increase of liver cancer cells to remain in the G1/S phase and to suppress growth. This treatment restored p21WAF1/Cip1, induced Beclin‐1, and repressed cyclin D1. In addition, sustained suppression of SIRT7 reduced the in vivo tumor growth rate in a mouse xenograft model. To explore mechanisms in SIRT7 regulation, microRNA (miRNA) profiling was carried out. This identified five significantly down‐regulated miRNAs in HCC. Bioinformatics analysis of target sites and ectopic expression in HCC cells showed that miR‐125a‐5p and miR‐125b suppressed SIRT7 and cyclin D1 expression and induced p21WAF1/Cip1‐dependent G1 cell cycle arrest. Furthermore, treatment of HCC cells with 5‐aza‐2′‐deoxycytidine or ectopic expression of wildtype but not mutated p53 restored miR‐125a‐5p and miR‐125b expression and inhibited tumor cell growth, suggesting their regulation by promoter methylation and p53 activity. To show the clinical significance of these findings, mutations in the DNA binding domain of p53 and promoter methylation of miR‐125b were investigated. Four out of nine patients with induced SIRT7 carried mutations in the p53 gene and one patient showed hypermethylation of the miR‐125b promoter region. Conclusion: Our findings suggest the oncogenic potential of SIRT7 in hepatocarcinogenesis. A regulatory loop is proposed whereby SIRT7 inhibits transcriptional activation of p21WAF1/Cip1 by way of repression of miR‐125a‐5p and miR‐125b. This makes SIRT7 a promising target in cancer therapy. (HEPATOLOGY 2013)

The JAK2 V617F mutation in de novo acute myelogenous leukemias

A missense somatic mutation in JAK2 gene (JAK2 V617F) has recently been reported in chronic myeloproliferative disorders, including polycythemia vera, essential thrombocythemia and myelofibrosis with myeloid metaplasia, strongly suggesting its role in the pathogenesis of myeloid disorders. As activation of JAK2 signaling is occurred in other malignancies as well, we have analysed 558 tissues from common human cancers, including colon, breast and lung carcinomas, and 143 acute adulthood leukemias by polymerase chain reaction – single strand conformation polymorphism analysis. We found three JAK2 mutations in the 113 acute myelogenous leukemias (AMLs) (2.7%), but none in other cancers. The mutations consisted of two V617F mutations and one K607N mutation. None of the AML patients with the JAK2 V617F mutation had a history of previous hematologic disorders. This is the first report on the JAK2 gene mutation in AML, and the data indicated that the JAK2 gene mutation may not only contribute to the development of chronic myeloid disorders, but also to some AMLs.

Somatic mutations of TRAIL-receptor 1 and TRAIL-receptor 2 genes in non-Hodgkin's lymphoma

Tumor necrosis factor-related apoptosis-inducing ligand-receptor 1 (TRAIL-R1) and tumor necrosis factor-related apoptosis-inducing ligand-receptor 2 (TRAIL-R2) are cell-surface receptors involved in tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced cell-death signaling. TRAIL-R1 and TRAIL-R2 genes have recently been mapped to chromosome 8p21-22, which is a frequent site of allelic deletions in many types of human tumors, including non-Hodgkins lymphoma (NHL). Because TRAIL/TRAIL receptor system plays an important role in lymphocyte homeostasis, we hypothesized that the mutations of TRAIL-R1 and TRAIL-R2 may be involved in the development of NHL and that such mutations may be responsible for the allelic losses of 8p21-22 in NHL. In this study, we analysed the entire coding region of TRAIL-R2 gene and the death domain region of TRAIL-R1 gene for the detection of the somatic mutations in a series of 117 human NHLs using polymerase chain reaction (PCR)-based single strand conformation polymorphism (SSCP) analysis. Overall, eight tumors (6.8%) were found to have two TRAIL-R1 gene mutations or six TRAIL-R2 gene mutations. Interestingly, of the eight mutations, six missense mutations (two TRAIL-R1 and four TRAIL-R2) were detected in the death domains and one nonsense mutation of TRAIL-R2 was detected just before the death domain. Our data suggest that somatic mutations of TRAIL-R1 and TRAIL-R2 genes may play a role in the pathogenesis of some NHLs and that TRAIL-R1 and TRAIL-R2 genes might be the relevant genes to the frequent loss of chromosome 8p21-22 in human NHL.

Molecular changes from dysplastic nodule to hepatocellular carcinoma through gene expression profiling

Progression of hepatocellular carcinoma (HCC) is a stepwise process that proceeds from pre‐neoplastic lesions—including low‐grade dysplastic nodules (LGDNs) and high‐grade dysplastic nodules (HGDNs)—to advanced HCC. The molecular changes associated with this progression are unclear, however, and the morphological cues thought to distinguish pre‐neoplastic lesions from well‐differentiated HCC are not universally accepted. To understand the multistep process of hepato‐carcinogenesis at the molecular level, we used oligo‐nucleotide microarrays to investigate the transcription profiles of 50 hepatocellular nodular lesions ranging from LGDNs to primary HCC (Edmondson grades 1‐3). We demonstrated that gene expression profiles can discriminate not only between dysplastic nodules and overt carcinoma but also between different histological grades of HCC via unsupervised hierarchical clustering with 10,376 genes. We identified 3,084 grade‐associated genes, correlated with tumor progression, using one‐way ANOVA and a one‐versus‐all unpooled t test. Functional assignment of these genes revealed discrete expression clusters representing grade‐dependent biological properties of HCC. Using both diagonal linear discriminant analysis and support vector machines, we identified 240 genes that could accurately classify tumors according to histological grade, especially when attempting to discriminate LGDNs, HGDNs, and grade 1 HCC. In conclusion, a clear molecular demarcation between dysplastic nodules and overt HCC exists. The progression from grade 1 through grade 3 HCC is associated with changes in gene expression consistent with plausible functional consequences. Supplementary material for this article can be found on the HEPATOLOGY website (http://www.interscience.wiley.com/jpages/0270‐9139/suppmat/index.html). (HEPATOLOGY 2005;42:809–818.)