Efficiency of serum copper/zinc ratio for differential diagnosis of patients with and without lung cancer

We examined serum copper (Cu), serum zinc (Zn), and the serum copper/zinc ratio (Cu/Zn) in 162 patients. All of them were seen to have an abnormal shadow in the chest X-ray films, that is, 109 patients with lung cancer (LC) and 53 patients with no lung cancer (NLC). The mean Cu and Cu/Zn in LC patients were significantly higher than those in NLC patients (p<0.05). In LC patients, Cu and Cu/Zn were higher and Zn was lower in advanced tumors than early ones. There was a significantly clear relation between Cu or Cu/Zn and the tumor (T) stages. When the relative risk (RR) of LC was estimated, it was seen that the higher Cu and Cu/Zn became, the higher RR became. Furthermore, we showed the sensitivity of the receiver operator characteristic of the test (ROC) curve for Cu, Cu/Zn, and carcinoembryonic antigen (CEA) to diagnose LC, as explained in a paragraph of methods.The determinations of Cu, Zn, and Cu/Zn are simple and inexpensive. They also appear to have a great diagnostic value in determining the local invasion of LC and as a screening test in the high-risk patients for LC.     

Characterization of a variant of carcinoembryonic antigen (CEA) using monoclonal and polyclonal anti-CEA antibodies

A variant of the carcinoembryonic antigen (CEA) with lower molecular weight than a CEA reference preparation has been separated from CEA. Using a polyclonal, spleen absorbed anti-CEA antiserum, the variant crossreacts with reference CEA in immunodiffusion. The CEA-activity of the variant has been demonstrated using an enzyme-immunoassay with monoclonal CEA specific antibodies. There is sufficient immunological evidence that this variant is a distinct antigen different from the crossreactive antigens described so far. The reactivity of the polyclonal anti-CEA antiserum with the CEA variant was abolished by absorption against the immobilized variant.     

Colon tumour secretopeptidome: Insights into endogenous proteolytic cleavage events in the colon tumour microenvironment

The secretopeptidome comprises endogenous peptides derived from proteins secreted into the tumour microenvironment through classical and non-classical secretion. This study characterised the low-M_r (< 3 kDa) component of the human colon tumour (LIM1215, LIM1863) secretopeptidome, as a first step towards gaining insights into extracellular proteolytic cleavage events in the tumour microenvironment. Based on two biological replicates, this secretopeptidome isolation strategy utilised differential centrifugal ultrafiltration in combination with analytical RP-HPLC and nanoLC-MS/MS. Secreted peptides were identified using a combination of Mascot and post-processing analyses including MSPro re-scoring, extended feature sets and Percolator, resulting in 474 protein identifications from 1228 peptides (≤ 1% q-value, ≤ 5% PEP) — a 36% increase in peptide identifications when compared with conventional Mascot (homology ionscore thresholding). In both colon tumour models, 122 identified peptides were derived from 41 cell surface protein ectodomains, 23 peptides (12 proteins) from regulated intramembrane proteolysis (RIP), and 12 peptides (9 proteins) generated from intracellular domain proteolysis. Further analyses using the protease/substrate database MEROPS, (http://merops.sanger.ac.uk/), revealed 335 (71%) proteins classified as originating from classical/non-classical secretion, or the cell membrane. Of these, peptides were identified from 42 substrates in MEROPS with defined protease cleavage sites, while peptides generated from a further 205 substrates were fragmented by hitherto unknown proteases. A salient finding was the identification of peptides from 88 classical/non-classical secreted substrates in MEROPS, implicated in tumour progression and angiogenesis (FGFBP1, PLXDC2), cell–cell recognition and signalling (DDR1, GPA33), and tumour invasiveness and metastasis (MACC1, SMAGP); the nature of the proteases responsible for these proteolytic events is unknown. To confirm reproducibility of peptide fragment abundance in this study, we report the identification of a specific cleaved peptide fragment in the secretopeptidome from the colon-specific GPA33 antigen in 4/14 human CRC models. This improved secretopeptidome isolation and characterisation strategy has extended our understanding of endogenous peptides generated through proteolysis of classical/non-classical secreted proteins, extracellular proteolytic processing of cell surface membrane proteins, and peptides generated through RIP. The novel peptide cleavage site information in this study provides a useful first step in detailing proteolytic cleavage associated with tumourigenesis and the extracellular environment. This article is part of a Special Issue entitled: An Updated Secretome.     

DNA vaccination against tumors

DNA vaccines have been used to generate protective immunity against tumors in a variety of experimental models. The favorite target antigens have been those that are frequently expressed by human tumors, such as carcinoembryonic antigen (CEA), ErbB2/neu, and melanoma-associated antigens. DNA vaccines have the advantage of being simple to construct, produce and deliver. They can activate all arms of the immune system, and allow substantial flexibility in modifying the type of immune response generated through codelivery of cytokine genes. DNA vaccines can be applied by intramuscular, dermal/epidermal, oral, respiratory and other routes, and pose relatively few safety concerns. Compared to other nucleic acid vectors, they are usually devoid of viral or bacterial antigens and can be designed to deliver only the target tumor antigen(s). This is likely to be important when priming a response against weak tumor antigens. DNA vaccines have been more effective in rodents than in larger mammals or humans. However, a large number of methods that might be applied clinically have been shown to ameliorate these vaccines. This includes in vivo electroporation, and/or inclusion of various immunostimulatory molecules, xenoantigens (or their epitopes), antigen-cytokine fusion genes, agents that improve antigen uptake or presentation, and molecules that activate innate immunity mechanisms. In addition, CpG motifs carried by plasmids can overcome the negative effects of regulatory T cells. There have been few studies in humans, but recent clinical trials suggest that plasmid/virus, or plasmid/antigen-adjuvant, prime-boost strategies generate strong immune responses, and confirm the usefulness of plasmid-based vaccination.     

抗癌胚抗原免疫毒素生物学功能的研究

为了研究免疫毒素发挥生物学作用的全过程,构建了两种不同形式的基于假单胞菌外毒素(Pseudomonas exotoxin A,PE)的抗癌胚抗原（carcinoembryonic antigen, CEA）免疫毒素CEA(Fv)/PE38/KDEL和PE35/CEA(Fv)/KDEL.用流式细胞术经间接免疫荧光法测定免疫毒素的抗原结合活性.免疫毒素的内化(internalization)通过间接免疫荧光标记的方法在激光共聚焦显微镜下进行检测,采用流式细胞术进行免疫毒素内化的定量分析.细胞凋亡采用FITC-annexin V/PI双荧光染色方法进行分析.用噻唑蓝(MTT)法测定免疫毒素的靶细胞杀伤功能.对两种形式的免疫毒素在各个功能阶段的性质进行研究和比较,全面展现了免疫毒素发挥生物学功能的过程及各个生物作用过程之间的相互影响,为详尽的机制研究和免疫毒素的进一步优化改造提供了重要的依据.     

Molecular Paths Linking Metabolic Diseases,Gut Microbiota Dysbiosis and Enterobacteria Infections

Alterations of both ecology and functions of gut microbiota are conspicuous traits of several inflammatory pathologies, notably metabolic diseases such as obesity and type 2 diabetes. Moreover, the proliferation of enterobacteria, subdominant members of the intestinal microbial ecosystem, has been shown to be favored by Western diet, the strongest inducer of both metabolic diseases and gut microbiota dysbiosis. The inner interdependence between the host and the gut microbiota is based on a plethora of molecular mechanisms by which host and intestinal microbes modify each other. Among these mechanisms are as follows: (i) the well-known metabolic impact of short chain fatty acids, produced by microbial fermentation of complex carbohydrates from plants; (ii) a mutual modulation of miRNAs expression, both on the eukaryotic (host) and prokaryotic (gut microbes) side; (iii) the production by enterobacteria of virulence factors such as the genotoxin colibactin, shown to alter the integrity of host genome and induce a senescence-like phenotype in vitro; (iv) the microbial excretion of outer-membrane vesicles, which, in addition to other functions, may act as a carrier for multiple molecules such as toxins to be delivered to target cells. In this review, I describe the major molecular mechanisms by which gut microbes exert their metabolic impact at a multi-organ level (the gut barrier being in the front line) and support the emerging triad of metabolic diseases, gut microbiota dysbiosis and enterobacteria infections.     

Differential patterns of expression of glycosylphosphatidylinositol-anchored carcinoembryonic antigen and alkaline phosphatase in various cancer cell lines

The expression of glycosylphosphatidylinositol (GPI-anchored) carcinoembryonic antigen (CEA) and alkaline phosphatase (ALP) on the cell surface of various cancer cell lines and a lung diploid cell line (WI38) was investigated, with exposure of the cell lines to a cell differentiation agent (sodium butyrate) to induce cell differentiation and expression of the two tumor-associated antigens. In three colon (SW1222, SW1116, and HT-29) and stomach (MKN-45) cancer cell lines, all of which are double producers of CEA and ALP, the maximum expression of GPI-anchored CEA occurred with butyrate at a lower concentration than did that of GPI-anchored ALP. GPI-anchored ALP derived from colon (SW1222 and SW1116) and stomach (MKN-45 and MKN-1) cancer cell lines was heat-stable with and without exposure to butyrate, but GPI-anchored ALP derived from lung cancer cell lines (PC-6, PC13, PC-14, WI26VA4, and WI38VA13) showed a variety of heat stabilities, depending on cell line, butyrate exposure, and SV40 transformation. This revised version was published online in July 2006 with corrections to the Cover Date.     

癌胚抗原(CEA)启动子在肿瘤细胞中的特异表达及介异bak基因诱发凋亡

构建由癌胚抗原 (CEA)启动子控制报道基因增强型绿色荧光蛋白 (EGFP)表达的重组表达质粒pCEA EGFP .用转染细胞后检测荧光的方法对CEA阳性细胞进行简便、直观的检测 ,并结合流式细胞计数对CEA启动子在人结直肠腺癌细胞LS 1 74T、结肠癌细胞SW 4 80、肺腺癌细胞A5 49、人宫颈癌细胞HeLa和人喉癌细胞HEp 2中的活性进行了分析 ,发现其在SW4 80、LS 1 74T、A5 49中活性较强 ,而在HeLa和HEp 2中无活性 .构建由CEA启动子控制凋亡基因bak表达的重组表达质粒pCEA bak ,转染HeLa及SW 4 80细胞 ,用Hoechst332 5 8染色及PI染色 流式细胞计数分析的方法证明 ,pCEA bak转染能够特异性引起SW 4 80细胞的凋亡 .结果表明 ,CEA启动子具有很好的特异性 ,CEA介导bak基因的方法可望用于CEA阳性癌细胞的靶向性基因治疗 .