Reduced Apaf-1 expression in human cutaneous melanomas

Malignant melanoma is a life-threatening skin cancer due to its highly metastatic character and resistance to radio- and chemotherapy. It is believed that the ability to evade apoptosis is the key mechanism for the rapid growth of cancer cells. However, the exact mechanism for failure in the apoptotic pathway in melanoma cells is unclear. p53, the most frequently mutated tumour suppressor gene in human cancers, is a key apoptosis inducer. However, p53 mutation is only found in 15-20% of melanoma biopsies. Recently, it was found that Apaf-1, a downstream target of p53, is inactivated in metastatic melanoma. Specifically, loss of heterozygosity (LOH) of the Apaf-1 gene was found in 40% of metastatic melanoma. To determine if loss of Apaf-1 expression is indeed involved in melanoma progression, we employed the tissue microarray technology and examined Apaf-1 expression in 70 human primary malignant melanoma biopsies by immunohistochemistry. Our data showed that Apaf-1 expression is significantly reduced in melanoma cells compared with normal nevi (chi(2)=6.02, P=0.014). Our results also revealed that loss of Apaf-1 was not associated with the tumour thickness, ulceration or subtype, patient's gender, age and 5-year survival. In addition, our in vitro apoptosis assay revealed that overexpression of Apaf-1 can sensitise melanoma cells to anticancer drug treatment. Taken together, our data indicate that Apaf-1 expression is significantly reduced in human melanoma and that Apaf-1 may serve as a therapeutic target in melanoma.     

CDH22 expression is reduced in metastatic melanoma

Cadherin-like protein 22 (CDH22) is a transmembrane glycoprotein implicated in cell-cell adhesion and cancer metastasis. The expression of CDH22 has been shown to be increased in colorectal cancers. However, the role of CDH22 in melanomagenesis is not known. To investigate the role of CDH22 in melanoma progression, we examined the expression of CDH22 in melanocytic lesions at different stages and analysed the correlation between CDH22 expression and clinicopathlogic parameters and patient survival. Using tissue microarray and immunohisto-chemistry, we evaluated CDH22 staining in 76 dysplastic nevi, 247 primary melanomas, and 143 metastatic melanomas. We found that metastatic melanomas had a significantly higher percentage of negative CDH22 staining than dysplastic nevi (P = 0.012) and primary melanomas (P = 0.038). CDH22 expression was also reduced in thick (≥2 mm) and ulcerative melanomas (P = 0.003 and 0.022, respectively). Melanomas of AJCC stage II, III, and IV had a higher percentage of negative CDH22 staining than AJCC stage I melanomas (P = 0.004, P < 0.0001, and P = 0.009, respectively). Melanomas with negative CDH22 expression had significantly poorer disease-specific 5-year survival than those with positive CDH22 staining. Additionally, CDH22 expression depended on AJCC stage to predict patient survival. These data indicate that reduced CDH22 expression is associated with melanoma metastasis and poor patient prognosis.     

PUMA expression is significantly reduced in human cutaneous melanomas

Cutaneous malignant melanoma is an aggressive form of skin cancer, characterized by strong chemoresistance and poor patient prognosis. The molecular mechanisms underlying its resistance to chemotherapy remain unclear but are speculated to involve the dysregulation of apoptotic pathways. In this study, we sought to determine whether PUMA (p53 upregulated modulator of apoptosis) contributes to human melanoma formation, tumor progression, and survival. We used tissue microarray and immunohistochemistry to examine PUMA expression in 107 primary melanomas, 51 metastatic melanomas, and 64 dysplastic nevi. Here we report that PUMA expression is significantly weaker in primary melanomas compared to dysplastic nevi (P<0.0001), and is further reduced in metastatic melanomas compared to primary tumors (P=0.001). We show that weak PUMA expression in melanoma correlates with poorer overall and disease-specific 5-year survival (P<0.005 and P<0.001, respectively) of melanoma patients and that PUMA expression in tumor tissue is an independent predictor of both overall and disease-specific 5-year survival (P=0.05). Additionally, we show that exogenous PUMA expression in human melanoma cell lines (both wild type and mutant p53) results in significant apoptotic cell death. Our results suggest that PUMA expression may be an important prognostic marker for human melanoma and that adenoviral delivery of PUMA sensitizes melanoma cells to apoptosis.     

No ING1 mutations in human brain tumours but reduced expression in high malignancy grades of astrocytoma

The ING1 family of proteins has been shown to have regulatory functions in oncogenesis, apoptosis, DNA repair and cell cycle regulation. Here we present the first report on LOH analysis of the ING1 locus, mutation analysis of the complete coding sequence including intron-exon boundaries and expression analysis of the different ING1 splice products and protein isoforms in primary brain tumours. No somatic ING1 mutations were detected. Semi-quantitative analysis revealed higher levels of p33ING1b RNA in benign than in malignant lesions. This correlation was significant in a subset of 37 astrocytic tumours WHO grades I to IV. ING1 protein isoforms p47ING1a, p33ING1b and p24ING1c were found to be expressed variably in this series. Our findings support a regulatory contribution of ING1 to the development or progression of brain tumours.     

胰腺癌中p33ING1b基因变化及其表达研究

目的检测胰腺癌中p33ING1b蛋白表达及基因变化情况,以了解p33ING1b在肿瘤发生过程中的作用.方法采用免疫组化、聚合酶链反应单链构象多态性技术(PCR-SSCP)和杂合型缺失(LOH)技术,检测胰腺癌中p33ING1b表达及p33ING1b基因变化情况.结果ING1b蛋白的阳性表达率为85%(34/40),SSCP显示在40例病例中仅1例突变,LOH率为60.9%(14/23).结论突变与缺失表达并不是p33ING1b的主要失活形式,染色体改变可能导致p33ING1b功能下降,从而引发肿瘤发生.     

Correlation of VEGF and COX-2 Expression with VM in Malignant Melanomas

OBJECTIVE To investigate the relationship between vascular epithelial growth factor (VEGF) and cyclooxygenase-2 (COX-2) in melanomas and the expressive difference of VEGF and COX-2 between melanomas with and without vasculogenic mimicry(VM).METHODS Sixty cases of malignant melanomas emoeaaea In paraffin were studied. The tumors were divided into a high-grade malignant group and a low-grade malignant group based on their tumor type, atypia and survival time of the patient. Then tissue microarrays were produced from these paraffin-embedded tumor tissues which were stained for VEGF, COX-2 and PAS. The difference in expression between VEGF and COX-2 in the malignant melanomas was compared using a grid-count. In addition, the tumors were also divided into mimicry and non-mimicry groups based on their PAS staining. Then the differences between the PAS positive and negative areas of the 2 groups were compared.RESULTS In malignant melanomas with VM, VEGF and COX-2 expression was less in tumors in which VM was absent, but VEGF, COX-2 expression in high-grade malignant melanomas was higher than that in low-grade grade malignant melanomas. Expression of VEGF was correlated with COX-2 expression.CONCLUSION VM exists in some high-grade malignant melanomas. The differences and relations between VEGF and COX-2 showed that some high-grade malignant melanomas possess a unique molecular-mechanism of tumor metastasis and blood supply.     

Gene expression profiling identifies tumour markers potentially playing a role in uveal melanoma development

Microarray is a powerful tool to compare the gene expression of different tumour specimens and cell lines simultaneously and quantitatively. To get a better insight into genes that are involved in uveal melanoma tumorigenesis, we compared the gene expression profiles of 12 different uveal melanoma cell lines with three melanocyte cell cultures obtained from healthy donor eyes. Gene expression profiles were obtained by nylon filter arrays, containing 1176 gene spots related to cancer development. The expression levels of selected genes were validated on cell lines and primary uveal melanomas by real time RT-PCR, and were subsequently included in cluster analysis. Four candidate tumour markers, Laminin Receptor 1, Endothelin 2, Von Hippel Lindau Binding protein 1 and Cullin 2, have been selected from genes that were differentially expressed in the uveal melanoma cell lines compared to the normal uveal melanocytes. In primary uveal melanomas, these four markers could discriminate between two classes of uveal melanoma, which may be indicative of a differential disease process.     

抑癌基因ING3在大肠癌中表达及意义

目的：探讨ING3在大肠癌中的表达及意义。方法收集82例大肠癌临床手术标本,通过荧光定量PCR检测癌旁正常黏膜中与癌组织中ING3的表达水平,分析ING3的表达水平与临床病理特征及预后的关系。结果51％的大肠癌患者中,ING3 mRNA在癌组织中的表达水平显著低于癌旁组织, ING3的表达与临床T分期、M 分期及TNM分期密切相关（P＜0．05）。单因素分析显示出T、N、M分期、TNM分期及ING3的表达均与患者术后总无病生存率相关（ P＜0．05）。 ING3低表达组患者长期无病生存率较中高表达组患者显著降低（ P＜0．05）。结论 ING3的表达与大肠癌发生发展相关,有望成为大肠癌早期诊断早期治疗的分子靶点。     

ING5在大肠癌中的表达及意义

目的:为研究大肠癌发生发展分子机制,检测分析ING5在大肠癌中的表达趋势及意义。方法:荧光定量PCR法检测临床手术标本中ING5信使RNA的表达,统计分析基因表达与临床病理特征之间的关系,分析影响预后因素。结果:ING5在大肠癌中呈低表达,ING5的表达与病理T、N、M分级及临床TNM分期密切相关,ING5低表达组患者长期无病生存率显著降低。结论:ING5的表达情况与患者无病生存相关并且可作为影响大肠癌预后的一个独立影响因素。ING5与大肠癌的发生发展密切相关。     

肺癌中ING1基因变化及其表达水平的研究

目的：检测肺癌中抑癌基因ING1微卫星杂合性缺失（LOH）及其主要蛋白产物p33^ING1b的表达情况，以探讨ING1基因改变-9肺癌发生发展的关系。方法：采用银染PCR—SSCP法检测肺癌组织ING1基因微卫星LOH发生的频率；应用组织芯片技术和免疫组化方法，检测肺癌p33^ING1b蛋白表达水平。结果：70例肺癌组织ING1基因4个微卫星位点的总杂合性缺失率为55．7％（39／70），并且越靠近ING1基因位点，发生率越高，但与临床病理参数无关。217例肺癌p33^ING1b蛋白的LOH发生率为47．0％（102／217），鳞状细胞癌高于腺癌、腺鳞癌和细支气管肺泡癌（P〈0．05），其余类型间差异无显著性（P〉0．05）；p33^ING1b的表达与其他临床病理参数不相关（P〉0．05），但与LOH发生频率相关（P〈0．05）。结论：p33^ING1b蛋白在肺癌组织中存在高频率的低表达或失表达，并且ING1基因的微卫星LOH发生频率也很高。推测LOH是该基因异常表达的重要原因，其结果可能导致该基因的下调和（或）蛋白质功能的失活，从而丧失其对细胞的生长抑制作用，促进了肿瘤的发生。     

ＩＮＧ３基因与肿瘤关系的研究进展

ＩＮＧ基因家族（ＩＮＧ１ ５）被认为是２型肿瘤抑制基因,参与细胞生长、增殖、细胞周期调控和细胞衰老等不同的生物学过程。新近发现的ＩＮＧ３基因,定位于染色体７ｑ３１.３,广泛表达于正常人体组织。但在多种类型肿瘤中ＩＮＧ３的表达下调或缺失,例如大肠癌、头颈癌以及皮肤恶性黑色素瘤等。该现象表明,多种机制导致的ＩＮＧ３失活可能促进了肿瘤的发生。本文对ＩＮＧ３基因与不同类型肿瘤关系的研究进展作一综述。     

Prognostic significance of RUNX3 expression in human melanoma

BACKGROUND:

RUNX3 is a tumor suppressor that plays important roles in cell proliferation, apoptosis, and metastasis. The authors investigated the role of RUNX3 in melanoma pathogenesis and analyzed the prognostic impact of RUNX3 expression in a large series of melanoma patients.

METHODS:

Two sets of tissue microarrays were constructed, including 440 cases of melanomas (202 for the training set and 238 for the validation set) and 88 cases of nevi (25 normal nevi and 63 dysplastic nevi). RUNX3 expression was evaluated by immunohistochemistry.

RESULTS:

Positive RUNX3 expression was observed in 56%, 54%, 33%, and 24% of the biopsies in normal nevi, dysplastic nevi, primary melanoma, and melanoma metastases, respectively. Significant differences for positive nuclear RUNX3 staining were observed between dysplastic nevi and primary melanomas (P = .002, chi‐square test), between dysplastic nevi and melanoma metastases (P < .001, chi‐square test), and between primary melanoma and melanoma metastases (P = .045, chi‐square test). Loss of RUNX3 expression was correlated with a worse 5‐year survival of melanoma patients in both training and validation sets. Furthermore, loss of RUNX3 expression was also correlated with a poor 5‐year disease‐specific survival in primary melanoma (P = .001) and metastatic melanoma patients (P = .008). Multivariate Cox regression analysis revealed that positive RUNX3 expression is an independent prognostic factor to predict melanoma patient outcome.

CONCLUSIONS:

Our findings indicate that RUNX3 plays an important role in melanoma pathogenesis and may serve as a promising prognostic marker for melanoma. Cancer 2011;. © 2010 American Cancer Society.     

The new tumor suppressor genes ING: genomic structure and status in cancer

The Inhibitor of Growth 1 (ING1) gene has been identified and characterized as a Type-II tumor suppressor gene (TSG). Subsequently, 4 additional members of the family were identified by homology search. ING proteins contain a nuclear localization sequence (NLS) and a plant homeo domain (PHD) finger motif in their C-terminus. These proteins are involved in numerous signaling pathways especially in 2 tumor suppressor pathways: apoptosis and senescence. In human tumors, several studies have shown that the expression of ING1 is frequently lost or downregulated. It occurs most frequently at the RNA level, and thus epigenetics mechanism could be involved. We summarize the current knowledge on ING proteins functions and their involvement in various signaling pathways. We also review the studies that have investigated the ING protein status in human tumors. The interest of ING proteins as biomarkers and their role in tumor initiation and progression is discussed.     

ING2 is upregulated in colon cancer and increases invasion by enhanced MMP13 expression

Inhibitor of growth 2 (ING2) is associated with chromatin remodeling and regulation of gene expression by binding to a methylated histone H3K4 residue and recruiting HDAC complexes to the region. The aim of our study is to investigate the regulation of ING2 expression and the clinical significance of upregulated ING2 in colon cancer. Here, we show that the ING2 mRNA level in colon cancer tissue increased to more than twice than that in normal mucosa in the 45% of colorectal cancer cases that we examined. A putative NF‐κB binding site was found in the ING2 promoter region. We confirmed that NF‐κB could bind to the ING2 promoter by EMSA and luciferase assays. Subsequent microarray analyses revealed that ING2 upregulates expression of matrix metalloproteinase 13 (MMP13), which enhances cancer invasion and metastasis. ING2 regulation of MMP13 expression was confirmed in both ING2 overexpression and knock down experiments. MMP13 expression was further induced by coexpression of ING2 with HDAC1 or with mSin3A, suggesting that the ING2‐HDAC1‐mSin3A complex members regulates expression of MMP13. In vitro invasion assay was performed to determine functional significance of ING2 upregulation. ING2 overexpressed cells exhibited greater invasive potential. Taken together, upregulation of ING2 was associated with colon cancer and MMP13‐dependent cellular invasion, indicating that ING2 expression might be involved with cancer invasion and metastasis. Published 2009 UICC.     

ING4及SKP2在不同级别脑胶质瘤组织中的表达

[目的]研究抑癌基因ING4及癌基因SKP2在正常脑组织及不同级别胶质瘤组织中的表达．以及其表达水平与脑胶质瘤生物学特征的关系。[方法]采用免疫组织化学技术检测70例手术切除的不同级别胶质瘤组织人及正常脑组织中ING4及SKP2的表达情况。[结果]ING4主要表达于细胞核和／或浆，正常脑组织中ING4的表达明显高于低级别与高级别胶质瘤，差异显著（t=11．8及22．5，P〈0．05）；低级别胶质瘤组织中ING4的表达明显高于高级别胶质瘤，差异显著（t=-17．9，P〈0．05）。SKP2主要表达于细胞核，部分为胞浆，正常脑组织中SKP2的表达明显低于低级别与高级别胶质瘤，差异显著（t=13．7及29．5，P〈0．05）；低级别胶质瘤组织中SKP2的表达明显低于高级别胶质瘤，差异显著（t=16．5，P〈0．05）。[结论]ING4基因在胶质瘤组织中表达明显下调，且ING4表达下调与胶质瘤的分级有密切关系；SKP2基因在胶质瘤组织中表达显著上调，且SKP2表达上调与胶质瘤的分级有密切关系。     

Gene expression array profile of human osteosarcoma

The aims of this pilot study were to determine whether needle and open biopsies from osteosarcoma (OS) provide sufficient quality of mRNA for cDNA array analyses to gain insights into the expression profile of OS. A total of 22 samples collected from OS were used for array analyses. A primary cell culture was also established from one of the OS biopsies. Total RNA was extracted and probes were generated for cDNA arrays. cDNA probes were made for all the 22 samples. Two of these samples were needle core bone biopsies. Statistical analysis confirmed the reliability of array data obtained in 16 of the 22 samples. Known genes involved in bone metabolism and osteosarcoma were identified as highly expressed, and the putative new marker Ezrin was also identified. Confirmatory immunohistochemical staining using the Ezrin antibody was performed in a selection of samples.