Amygdalin-mediated inhibition of non-small cell lung cancer cell invasion in vitro

Lung cancer is a common malignant tumor claiming the highest fatality worldwide for a long period of time. Unfortunately, most of the current treatment methods are still based on the characteristics of cancer cells in the primary lesion and the prognosis is often much poorer in patients with metastatic cancers. Amygdalin, a natural product of glycosides and lots of evidence shows that amygdalin can inhibit the proliferation of some kinds of cancer cells. In this study, we first obtained the highly metastatic NSCLC cell lines H1299/M and PA/M and further treated these cells with amygdalin. We found that the in vitro proliferability of H1299/M and PA/M was inhibited, but such inhibition required higher concentration of amygdalin. When lower concentration of amygdalin was used for the experiments, we observed that the in vitro invasive and migration capacities of H1299/M and PA/M were significantly inhibited. These results strongly suggested that amygdalin was likely to have anti-metastatic NSCLC effect. This study offers information of the role of amygdalin that may be useful as a therapeutic target in lung tumors.     

Genetic variants in the KDM6B gene are associated with neurodevelopmental delays and dysmorphic features

Lysine‐specific demethylase 6B (KDM6B) demethylates trimethylated lysine‐27 on histone H3. The methylation and demethylation of histone proteins affects gene expression during development. Pathogenic alterations in histone lysine methylation and demethylation genes have been associated with multiple neurodevelopmental disorders. We have identified a number of de novo alterations in the KDM6B gene via whole exome sequencing (WES) in a cohort of 12 unrelated patients with developmental delay, intellectual disability, dysmorphic facial features, and other clinical findings. Our findings will allow for further investigation in to the role of the KDM6B gene in human neurodevelopmental disorders.     

JMJD2B/KDM4B inactivation in adipose tissues accelerates obesity and systemic metabolic abnormalities

Obesity is a serious global health issue; however, the roles of genetics and epigenetics in the onset and progression of obesity are still not completely understood. The aim of this study was to determine the role of Kdm4b, which belongs to a subfamily of histone demethylases, in adipogenesis and fat metabolism in vivo. We established conditional Kdm4b knockout mice. Inactivation of Kdm4b in adipocytes (K4bKO) induced profound obesity in mice on a high fat diet (HFD). The HFD‐fed K4bKO mice exhibited an increased volume of fat mass and higher expression levels of adipogenesis‐related genes. In contrast, the genes involved in energy expenditure and mitochondrial functions were down‐regulated. Supporting these findings, the energy expenditure of Kdm4b‐deficient cells was markedly decreased. In addition, progression of glucose intolerance and hepatic steatosis with hepatocellular damages was observed. These data indicate that Kdm4b is a critical regulator of systemic metabolism via enhancing energy expenditure in adipocytes.     

Prognostic value of FHIT, CTNNB1, and MUC1 expression in non-small cell lung cancer

Comprehensive expression analysis using microarrays has identified a number of differentially expressed genes in smoke-exposed bronchial epithelium and non-small cell lung cancers (NSCLCs). To evaluate the prognostic relevance of these proteins in NSCLCs, we used immunohistochemistry to investigate the expression of beta-catenin (CTNNB1), dickkopf, Xenopus, homolog of 3 (DKK3 gene), fibroblast growth factor receptor 3 (FGFR3), fragile histidine triad (FHIT), tumor protein p53 (TP53), mucin1 (MUC1), topoisomerase II alpha (TOP2A), and glutathione S-transferase-Pi (GST) in a cohort of patients (n = 125). We correlated the expression data with clinicopathologic features and clinical outcome. In addition, SNaPshot multiplex assays (Applied Biosystems, Darmstadt, Germany) and restriction fragment length polymorphism analysis were used to screen for activating point mutations at the hot spots of FGFR3 in a cohort of 30 samples of NSCLC. Using Kaplan-Meier analysis, we observed significantly better overall survival in adenocarcinomas compared with squamous cell cancers (P = .049). Loss of FHIT expression showed a strong association with shorter overall survival in both histologic types of NSCLC (squamous cell cancers, P < .001; adenocarcinomas, P = .001). In adenocarcinomas, the cytoplasmic expression of beta-catenin was associated with shorter survival (P = .012); MUC1 expression was associated with worse prognosis in patients with squamous cell cancers (P = .002). The nuclear staining of TP53 (P = .008) and TOP2A (P = .059) was associated with cancers without lymphonodal metastases. A correlation with positive staining of TOP2A (P = .03) and FGFR3 positivity (P = .057) was found in adenocarcinomas of male patients. Positive MUC1 stainings were associated with squamous cell cancers of male patients (P = .03). DKK3 expression did not show any significant association with clinical outcome or pathologic features. The screening of the FGFR3 sequence in lung cancers showed only wild-type sequences and did not detect mutations in the known hot spots for FGFR3 mutations. We conclude that the immunohistochemical loss of FHIT expression and the positivity for beta-catenin and MUC1 in NSCLC are useful prognostic markers, whereas the variable expression of TP53, TOP2A, and FGFR3 in relation to the different histologic types of NSCLC and sex of the patients is suggestive for different underlying molecular pathways.     

A molecular threading mechanism underlies Jumonji lysine demethylase KDM2A regulation of methylated H3K36

The dynamic reversible methylation of lysine residues on histone proteins is central to chromatin biology. Key components are demethylase enzymes, which remove methyl moieties from lysine residues. KDM2A, a member of the Jumonji C domain-containing histone lysine demethylase family, specifically targets lower methylation states of H3K36. Here, structural studies reveal that H3K36 specificity for KDM2A is mediated by the U-shaped threading of the H3K36 peptide through a catalytic groove within KDM2A. The side chain of methylated K36 inserts into the catalytic pocket occupied by Ni²⁺ and cofactor, where it is positioned and oriented for demethylation. Key residues contributing to K36me specificity on histone H3 are G33 and G34 (positioned within a narrow channel), P38 (a turn residue), and Y41 (inserts into its own pocket). Given that KDM2A was found to also bind the H3K36me3 peptide, we postulate that steric constraints could prevent α-ketoglutarate from undergoing an “off-line”-to-“in-line” transition necessary for the demethylation reaction. Furthermore, structure-guided substitutions of residues in the KDM2A catalytic pocket abrogate KDM2A-mediated functions important for suppression of cancer cell phenotypes. Together, our results deduce insights into the molecular basis underlying KDM2A regulation of the biologically important methylated H3K36 mark.     

KDM5B基因在乳腺癌中的表达及其临床意义

目的:研究KDM5B基因在乳腺癌组织中的表达情况,并探讨其表达差异与患者临床病理资料和预后的关系。方法:从肿瘤基因组图谱(TCGA)数据库中收集113例正常乳腺组织和1 090例乳腺癌数据集,下载KDM5B mRNA表达谱资料;收集中山大学附属第三医院甲乳外科2015年～2016年乳腺癌切除标本共90例,采用real-time PCR的方法检测乳腺癌及其癌旁组织中KDM5B mRNA的表达量,取中位数分为高表达组与低表达组,分析其与临床资料及病例特征的关系;利用Kaplan-Meier plotter分析KDM5B mRNA表达与乳腺癌患者预后的相关性。结果:KDM5B在乳腺癌中的表达均显著高于正常乳腺组织(P 0. 01)。在TCGA的乳腺癌数据中,KDM5B的表达与人表皮生长因子受体2(HER2)、雌激素受体(ER)、年龄、病理组织类型及淋巴结转移显著相关(P 0. 01),与孕激素受体(PR)、绝经及远处转移无显著相关。在我们收集的乳腺癌样本中,KDM5B的表达与HER2、年龄及淋巴结转移显著相关(P 0. 05),而与ER、PR、绝经、病理组织类型和远处转移无显著相关。KDM5B表达越高,乳腺癌患者总生存时间(HR=1. 39,P=0. 005)和无病生存时间(HR=1. 32,P 0. 001)越短。结论:在乳腺癌组织中KDM5B高表达,且与患者的预后相关; KDM5B的表达与乳腺癌患者HER2表达、年龄和淋巴结转移显著相关。     

EML4-ALK fusion gene in Korean non-small cell lung cancer

A fusion gene between echinoderm microtubule-associated protein-like 4 (EML4) and the anaplastic lymphoma kinase (ALK) has been identified in non-small cell lung cancers (NSCLCs). Although a few studies have evaluated EML4-ALK fusion genes in Korean NSCLCs, the prevalence of different EML4-ALK fusion variants has yet to be clearly assessed. Herein, we have examined the profiles of EML4-ALK fusion gene variants in Korean patients of NSCLCs. EML4-ALK fusion genes have been detected in 10 (6.0%) of 167 patients of NSCLCs and in 9 (7.4%) of 121 patients of adenocarcinoma. Of the 10 patients with fusion genes identified, 8 (80%) were E13;A20 (variant 1) and 2 (20%) were E6;A20, with an additional 33-bp sequence derived from intron 6 of EML4 (variant 3b). These results indicate that the profiles of EML4-ALK fusion gene variants in Korean patients of NSCLC may differ from those in other ethnic populations. Herein, we describe for the first time the profiles of EML4-ALK fusion variants of Korean patients with NSCLCs.     

Knockdown of KDM1B inhibits cell proliferation and induces apoptosis of pancreatic cancer cells

Pancreatic cancer (PC) is one of the common malignant tumors in digestive tract with a high fatality rate. The oncogenic role of lysine-specific demethylase1 (LSD1/KDM1?A) has been well recognized in PC. While, the role of its homolog LSD2 (KDM1B) in regulating PC progression is poorly understood. In this study, we attempted to evaluate the functional role of KDM1B in PC cells. The expression of KDM1B was detected by immunohistochemistry and immunoblotting in PC tissues and cells. Lentivirus-mediated shRNA was applied to silence KDM1B in PANC-1 and SW1990 cells. Cell proliferation was measured by MTT and Celigo assay. Cell apoptosis was determined by both Caspase-Glo^®3/7 assay and Flow cytometry. Intracellular signaling molecules were detected using a PathScan intracellular signaling array kit. In this study, we found KDM1B was highly expressed in PC tissues compared to paracancerous tissues. Moreover, elevated expression of KDM1B was detected in PC cell lines (BxPC-3, CFPAC-1, PANC-1 and SW1990) as compared with a normal human pancreatic duct epithelial cell line (HPDE6-C7). Further investigations revealed that KDM1B knockdown significantly inhibited PC cell proliferation. Furthermore, the apoptosis of PANC-1 and SW1990 cells was significantly increased after KDM1B knockdown. Notably, the activations of p-ERK1/2, p-Smad2, p-p53, cleaved PARP, cleaved Caspase-3, cleaved Caspase-7, p-eIF2a and Survivin were promoted by KDM1B knockdown, while IkBa was suppressed. Taken together, our findings provided new insights into the critical and multifaceted roles of KDM1B in the regulation of cell proliferation and apoptosis, and offered a potentially novel target in preventing the progression of PC.     

hnRNP A2/B1 mRNA在非小细胞肺癌和转移淋巴结中表达及意义

目的：探讨肺癌早期诊断基因核内不均一核糖体表达对肺癌淋巴结微转移的作用。观察hnRNP A2/B1 在各组织类型肺癌中的表达。方法：42例非小细胞肺癌患者开胸手术纵隔淋巴结清扫获取原发灶及纵隔淋巴结病理标本,12例肺良性病变手术切除标本作正常对照。荧光定量PCR检测切除肿块和淋巴结的hnRNP A2/B1 mRNA表达。结果：(1)转移性淋巴结中hnRNP A2/B1 mRNA 表达水平561.393显著高于无转移淋巴结156.669 (P＜0.05)。(2)Ⅲ期肺癌组织中hnRNP A2/B1 mRNA 表达水平305（249,533）显著高于Ⅰ期和Ⅱ期肺癌组织(2＝13.453,P＜0.01)。(3)hnRNP A2/B1 mRNA在鳞癌和腺癌中的表达水平分别为207(152,305)和249(177,476), Z=-0.899,P>0.05。(4)58组淋巴结阳性,84组阴性,42个病人142组区域淋巴结hnRNP A2/B1 mRNA阳性率为40.84%(58/142),12例肺良性疾病组织均阴性,HE染色42个病人的50组转移淋巴结阳性率为35.21%,与FQ-PCR 58枚（52.11%）无显著差异（P>0.05）。结论：NSCLC 癌组织存在着hnRNP A2/B1 的过度表达。NSCLC癌组织中hnRNP A2/B1表达水平与淋巴结转移状态及P-TNM 分期有关。 hnRNP A2/B1 检验敏感性、特异性高,可用于肺癌的辅助诊断的标记。     

Diverse ways to be specific: a novel Zn-binding domain confers substrate specificity to UTX/KDM6A histone H3 Lys 27 demethylase

Histone methylations are highly regulated by site-specific histone methyltransferases and demethylases. In this issue of Genes & Development, Sengoku and Yokoyama (pp. 2266-2277) demonstrate that a novel Zn-binding domain and the Jumonji domain of UTX/KDM6A (Lys demethylase 6A) recognize histone H3 and together function as a substrate specificity determinant for H3K27 demethylation. This study demonstrates the mechanism of site-specific demethylation by UTX/KDM6A and implicates that histone demethylases use diverse methods to accomplish target specificity.     

Cardiotoxicity of cisplatin-based chemotherapy in advanced non-small cell lung cancer patients

Cardiotoxicity is a well known consequence of cancer chemotherapy. Cisplatin-based combinations are standard regimens in the therapy of advanced non-small cell lung cancer. Administration of cisplatin-containing chemotherapy causes significant oxidative and nitrosative stress in some patients. Cardiac blood biomarkers can be used to evaluate cardiac status, may help to identify patients at risk myocardial damage evaluation and are able to detect subclinical, early-stage cisplatin-induced cardiotoxicity. The relevance of cardiovascular complications in cancer patients and identification of individual risk factors for developing cardiovascular toxicity merit further evaluation and a longer follow-up is needed.     

非小细胞型肺癌与弓形虫感染的关系

目的： 探讨非小细胞型肺癌（NSCLC）患者弓形虫(TOX)感染的状况。 方法: 用酶联免疫法(ELISA)检测168例NSCLC患者、同期住院的90例非肿瘤且无自身免疫性疾病患者以及90例健康体检者血中弓形虫循环抗原(TOX-CAg)、TOX-IgG及TOX-IgM抗体阳性率，比较各组间以及NSCLC组内性别、肿瘤分期和病理类型的弓形虫阳性率差异。 结果: 168例NSCLC患者血清中TOX-CAg阳性者34例(20.2%)，TOX-IgM阳性者36例(21.4%)，TOX-IgG阳性者27例(16.1%)；而90例其它疾病患者血清中3者分别为9例(10.0%)、10例(11.1%)和6例(6.7%)，90例健康组血清中3者分别为3例(3.3%)、4例(4.4%)和4例(4.4%)；NSCLC组与其它疾病组的TOX-CAg、IgG及IgM阳性率差异均有显著性(P<0.05)，NSCLC组与健康组的TOX-CAg、IgG及IgM差异更明显(P<0.01)，其它疾病组与健康组的TOX-CAg、IgG及IgM差异不明显(P>0.05)；另外，在肺癌分期方面，对于TOX-CAg和TOX-IgM，Ⅰ、Ⅱ期间无明显差异(P>0.05)，Ⅲ、Ⅳ期间也无明显差异(P>0.05)，但Ⅰ、Ⅱ期与Ⅲ期间差异显著（P<0.05），Ⅰ、Ⅱ期与Ⅳ期间差异显著（P<0.05或P<0.01），对于TOX-IgG抗体，各肿瘤分期间无明显差异(P>0.05)；肺腺、鳞癌与其它病理类型间无明显差异(P>0.05)；男女间无明显差异（P>0.05）。 结论: 非小细胞型肺癌患者的弓形虫感染率明显高于其他人，而且晚期肺癌的弓形虫感染率高于早期肺癌。     

Sirtuin SIRT6 suppresses cell proliferation through inhibition of Twist1 expression in non-small cell lung cancer

SIRT6 is a member of the NAD⁺-dependent class III deacetylase sirtuin family. Current studies have revealed that SIRT6 plays important roles in the epigenetic regulation of genes expression and contribute to the proliferation, differentiation and apoptosis of cancer cells. However, the biological function of SIRT6 in lung cancer has not been elucidated. The present study showed that the mRNA and protein levels of SIRT6 were decreased in human non-small cell lung cancer (NSCLC) tissues and cell lines. MTT assay showed that overexpression of SIRT6 could inhibit the proliferation in NSCLC cells. In contrast, SIRT6 knockdown using small interfering RNA promoted NSCLC cells proliferation. On the molecular level, we found that SIRT6 inhibited the expression of Twist1 both at the mRNA and protein levels in NSCLC cells. Taken together, these results demonstrated for the first time that SIRT6 suppressed NSCLC cells proliferation via down-regulation of Twist1 expression and might provide novel therapeutic targets in the treatment of lung cancer.     

p21活化激酶4在非小细胞肺癌中的表达及其临床意义

目的:探讨p21活化激酶4(PAK4)在人非小细胞肺癌细胞系及人非小细胞肺癌组织中的表达及其临床意义。方法:采用Western blot及实时荧光定量PCR检测人支气管上皮(HBE)细胞、非小细胞肺癌细胞(A549、NCI-H520、NCI-H460和NCI-H596细胞)、20例新鲜非小细胞肺癌组织及相应的癌旁组织中PAK4的表达情况;采用免疫组化检测210例非小细胞肺癌组织PAK4的表达情况;采用Kaplan-Meier法评估非小细胞肺癌患者术后5年生存率;用Cox比例风险模型分析PAK4对患者预后的影响。结果:非小细胞肺癌细胞(A549、NCI-H520、NCI-H460和NCI-H596细胞)PAK4蛋白及mRNA表达均显著高于HBE细胞(P0.05);20例非小细胞肺癌组织中PAK4蛋白及mRNA表达高于癌旁组织PAK4蛋白及mRNA表达(P0.05);10例转移性非小细胞肺癌组织PAK4 mRNA表达显著高于10例原发性非小细胞肺癌组织(P0.05);免疫组化染色结果示210例非小细胞肺癌PAK4评分显著高于对应的癌旁组织。临床资料分析显示PAK4蛋白表达与非小细胞肺癌的分化程度、淋巴结转移、远处转移及临床分期有关(P0.05);PAK4高表达组患者5年生存率显著低于PAK4蛋白低表达组患者(logrank检验,P0.05),PAK4蛋白表达是非小细胞肺癌患者的独立预后因素。结论:PAK4蛋白高表达是非小细胞肺癌患者死亡的独立危险因素。     

Human phenotype caused by biallelic KDM4B frameshift variant

KDM4B (MIM*609765, NM_015015.3, formerly JMJD2B) encodes a histone demethylase and regulates gene expression via demethylation, mainly of H3K9 tri-methylation. Heterozygous KDM4B loss-of-function variants cause autosomal dominant intellectual developmental disorder 65 (MIM#619320), which is characterized by global developmental delay, intellectual disability, language and gross motor delays, structural brain anomalies, characteristic facial features, and clinodactyly. Although the majority of reported patients have de novo pathogenic variants, some patients inherit pathogenic variants from affected parents. To our knowledge, only 23 patients with heterozygous KDM4B variants have been reported to date, and there are no reports of patients with biallelic KDM4B pathogenic variants. Herein, we report a female patient with a biallelic KDM4B frameshift variant (NM_015015.3: c.1384_1394delinsGGG, p.(Leu462Glyfs*43)) located at exon 12 of 23 protein-coding exons, which is thought to be subject to nonsense-mediated mRNA decay and no protein production. She presented developmental and language delays and a hypotonic and characteristic face. The patient's phenotype was more obvious than that of her mother, who is heterozygous for the same variant. Although declining birth rate (embryonic lethality in male mice) in homozygous knockout mice has been demonstrated, our report suggests that homozygous KDM4B frameshift variants can be viable in humans at least female.     

KDMs参与恶性肿瘤细胞上皮间质转化

转移是导致肿瘤患者预后差的根本原因。上皮间质转化（ epithehal?mesenchymal transition, EMT）是调控胚胎发生和组织发育过程中细胞形态和功能改变的复杂程序。 EMT在肿瘤转移过程中也起重要作用。已知表观遗传修饰在调控基因表达中起重要作用,其中包括组蛋白甲基化。组蛋白去甲基化酶（ his?tone demethylase）可通过特异性的作用于赖氨酸单甲基化、双甲基化和三甲基化调控基因表达。越来越多的研究表明组蛋白赖氨酸去甲基化酶（ histone lysine demethylases, KDMs）在调控MET相关基因表达中其重要作用。文章就KDMs在人类恶性肿瘤细胞的EMT过程中的研究进展作一综述。     

人非小细胞肺癌细胞中端粒酶活性的检测与研究

目的 研究非小细胞肺癌细胞癌组织中端粒酶活性表达,探讨端粒酶生表达与非小细胞肺癌发生发展的关系。方法 收集经手术及病理证实的非小细胞肺癌４８例标本,１２例肺癌癌旁肺组织、７例非肿瘤病例所取正常肺组织作对照。用端粒酶检测试剂盒检测组织本端粒酶活性。结果 ７５．００％（３６／４８）非小细胞肺癌组织标本检出端粒酶活性,８．３３％（１／１２）癌旁肺组织检出端粒酶活笥,７例非肿瘤标本所取的正常肺组织均未检出