Impact of the CCR5 gene polymorphism on the survival of metastatic melanoma patients receiving immunotherapy

Purpose Chemokines influence both tumor progression and anti-tumor immune response. A 32-bp-deletion polymorphism in the chemokine receptor 5 gene (CCR5Δ32) has been shown to result in a non-functional protein. This study was aimed at evaluating the potential impact of this gene polymorphism on disease progression and treatment outcome in patients with melanoma. Patients and methods CCR5 genotyping was performed by PCR on DNA extracted from serum samples of 782 cutaneous melanoma patients with known disease history and long-term clinical follow-up. Genotypes were correlated with patient survival and types of treatment. Results Of 782 melanoma patients, 90 (11.5%) were heterozygous and 12 (1.5%) were homozygous for CCR5Δ32. Analyzing the complete cohort, the disease-specific survival from date of primary diagnosis was not influenced by CCR5 status. Similarly, no significant impact could be detected on the treatment outcome of stage III patients. In 139 stage IV patients receiving immunotherapy, CCR5Δ32 was associated with a decreased survival compared to patients not carrying the deletion (median 12.5 vs. 20.3 months, P = 0.029). Multivariate analysis revealed the CCR5 genotype as an independent factor impacting disease-specific survival in this patient population (P = 0.002), followed by gender (P = 0.019) and pathological classification of the primary (pT; P = 0.022). Conclusion The presence of the CCR5Δ32 polymorphism in patients with stage IV melanoma results in a decreased survival following immunotherapy and may help to select patients less likely to benefit from this type of treatment. Selma Ugurel and David Schrama have contributed equally to this work.     

重组棉铃虫病毒的外源基因向其他生物的转移

本文报道了基因工程棉铃虫核多角体病毒(HaSNPV-AaIT)的外源蝎子毒素基因(AaIT)是否会向环境中的植物病原微生物或捕食性天敌转移的实验结果。首先,于实验室内将重组病毒HaSNPV-AaIT与棉花黄萎病病菌(VerticilliumdahliaeLleb.)进行了长达90d的混合培养,在混合培养30d,60d和90d后分别提取棉花黄萎病病菌的基因组DNA,用AaIT基因作探针进行点杂交,结果显示无阳性信号。另外,从多次施用过重组病毒的棉花田中采集了120只龟纹瓢虫和七星瓢虫,利用健康蚜虫饲养3-4d,用碱解液处理瓢虫体表后,从处理液中可以检测到病毒DNA;但瓢虫体表经碱解液和Dnase处理后,从瓢虫体内提取的基因组DNA,用PCR和斑点杂交的方法,都没有检测到AaIT的序列存在。本研究的实验结果说明,基因工程病毒的外源基因向其它生物转移的可能性极低。     

The SNPlex genotyping system: a flexible and scalable platform for SNP genotyping.

We developed the SNPlex Genotyping System to address the need for accurate genotyping data, high sample throughput, study design flexibility, and cost efficiency. The system uses oligonucleotide ligation/polymerase chain reaction and capillary electrophoresis to analyze bi-allelic single nucleotide polymorphism genotypes. It is well suited for single nucleotide polymorphism genotyping efforts in which throughput and cost efficiency are essential. The SNPlex Genotyping System offers a high degree of flexibility and scalability, allowing the selection of custom-defined sets of SNPs for medium- to high-throughput genotyping projects. It is therefore suitable for a broad range of study designs. In this article we describe the principle and applications of the SNPlex Genotyping System, as well as a set of single nucleotide polymorphism selection tools and validated assay resources that accelerate the assay design process. We developed the control pool, an oligonucleotide ligation probe set for training and quality-control purposes, which interrogates 48 SNPs simultaneously. We present performance data from this control pool obtained by testing genomic DNA samples from 44 individuals. in addition, we present data from a study that analyzed 521 SNPs in 92 individuals. Combined, both studies show the SNPlex Genotyping system to have a 99.32% overall call rate, 99.95% precision, and 99.84% concordance with genotypes analyzed by TaqMan probe-based assays. The SNPlex Genotyping System is an efficient and reliable tool for a broad range of genotyping applications, supported by applications for study design, data analysis, and data management.     

Analysis of single nucleotide polymorphisms in the promoter region of interleukin-10 by denaturing high-performance liquid chromatography.

Interleukin-10 (IL10), an anti-inflammatory cytokine, has been implicated in a variety of immune- and inflammatory-related diseases. We investigated the following SNPs: -1082, -819, -592 in the promoter region of IL10 in a normal (control) population and selected diseases: breast cancer (BrCa), systemic lupus erythematosus (SLE), and B-cell chronic lymphocytic leukemia (B-CLL) by denaturing high-performance liquid chromatography (DHPLC) and found distinct genotype and haplotype patterns. DHPLC was performed using the Transgenomic WAVE instrument, a mutational discovery tool that allows for high throughout analysis of SNPs. The principle of DHPLC is based on separation of homo- and heteroduplex formation of individual polymerase chain reaction products at specific melting temperatures and set gradients. The melting temperature selected for each SNP was based on size and sequence of the polymerase chain reaction product (for -1082, 57 degrees C; for -819, 58 degrees C; and for -592, 59.2 degrees C). Before fragment mutational analysis, all samples were denatured at 95 degrees C and slowly reannealed to allow for reassociation of different strands. Heteroduplex samples were easily distinguished from homoduplex samples. In order to identify wild type from homozygous mutant, two homoduplex polymerase chain reaction samples had to be mixed together, denatured at 95 degrees C and reannealed. The homozygous mutant, when combined with wild type, displayed a double peak on chromatogram. Once distinct chromatograms were established for each of the SNPs and the nucleotide changes confirmed by sequencing, genotype and haplotype frequencies were tabulated for the groups studied.     

FMMU白化豚鼠线粒体DNA RFLP分析研究

目的研究FMMU白化豚鼠的mtDNA,并与花色豚鼠mtDNA进行多态性分析比较,以确定其独特的生物学特性是否与mtDNA相关。方法用碱变性法提取FMMU白化豚鼠以及花色豚鼠的mtDNA,并用AvaⅠ、BalⅠ等12种限制性内切酶进行酶切和限制性片段长度多态性分析。结果与结论FMMU白化豚鼠mtDNA和花色豚鼠mtDNA的相对分子质量相同,约为16.7×103;FMMU白化豚鼠与花色豚鼠两品系的mtDNA经AvaⅠ、BalⅠ等内切酶酶切后有3-8个酶切位点,酶切图谱完全相同,经RFLP分析FMMU白化豚鼠与花色豚鼠的mtDNA之间缺乏多态性。本实验没有发现FMMU白化豚鼠的独特的生物学特性与mtDNA相关。     

Identification of the Gene Whose Mutation Leads to the Appearance of Recessive Lethal Germlings in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

The data are presented on genetic and molecular-genetic analysis of a mutant from the collection of morphological insertion mutants of Arabidopsis thaliana we obtained earlier, which belongs to the phenotypic class of recessive lethal germlings. A nucleotide DNA sequence, 147 bp in size, was identified, which adheres to the left border area of T-DNA insertion. The site of localization of the insertion was determined using computer analysis.__________Translated from Ontogenez, Vol. 36, No. 3, 2005, pp. 222–224.Original Russian Text Copyright © 2005 by Ogarkova, Tomilov, Tomilova, Tarasov.     

我国禽脑脊髓炎病毒分离株全基因组的测定

测定了我国禽脑脊髓炎病毒(avian encephalomyelitis virus,AEV)分离株L2Z株的全基因组核苷酸序列.该病毒株的3′和5′非编码区核苷酸序列用3′和5′RACE(cDNA末端快速扩增)法获得.基因组全长为7 059个核苷酸残基,包括494个核苷酸残基的5′非编码区、6 402个核苷酸残基的开放阅读框和136个核苷酸残基的3′非编码区及poly(A)尾巴.与已发表的AEV疫苗株1 143的基因组序列比较发现,它们之间核苷酸和氨基酸的同源性分别为98%和97.6%.结构蛋白(VP1～VP4)中,主要宿主保护性免疫原蛋白VP1氨基酸之间差异较小.与小RNA病毒科其它病毒属相比,在非结构蛋白3D中,预测的8个RNA依赖性RNA聚合酶主要结构域中的4个高度保守.从而进一步确认了AEV的分子特性.     

Tree Climbing Strategies for Primate Ecological Studies

Primate ecological studies can benefit from accessing the canopy to estimate intra-tree and inter-tree variation in food availability and nutrient value, patch and subpatch depletion, foraging efficiency, as well as nest structure and nesting behaviors, parasitic transmission and predator detectability. We compare several ways to access the canopy and examine their suitability for studies of primates. Two of them—the Single Rope Technique and the Climbing Spur Method—allow people to safely access almost all kinds of trees, regardless of their size, height or shape. Modern climbing gear and contemporaneous safety protocols, derived from rock climbers, speleologists, and industrial arborists, are reliable and appropriate for primate ecological studies. Climbing gear is specialized and still expensive for students, but tree climbing can be dangerous during specific maneuvres. Consequently, formal training and preliminary experience are essential before attempting to collect data. We discuss the physics of falling, risk assessment associated with a fall, knots, gear and safety precautions. Finally, we propose a Tree Climbing Safety Protocol adapted for 2 climbing methods and primate field ecology. Researchers should be aware that climbing safety depends on their own judgment, which must be based on competent instruction, experience, and a realistic assessment of climbing ability. Therefore, the information we provide should be used only to supplement competent personal instruction and training in situ. Although most primate observations have been and will mostly be done from the ground in the future, canopy information complements the observations. Canopy data will add a significant new dimension to our knowledge of primates by providing strategic information otherwise unavailable.     

Phosphorylation of Nucleotide Molecules in Hydrothermal Environments

Phosphorylation of AMP into ADP and ATP, that can outrun their hydrolysis, was made possible in a simulated hydrothermal environment when trimetaphosphate was used as the phosphate source. The best yields of phosphorylated products were obtained when the reaction fluids whose temperature was set at about 100 degrees centigrade was injected into the cold environment maintained at 0 degree in a recycling manner. Hydrothermal environments in the primitive ocean could also have served as prebiotic sites for phosphorylation, among others.     

Comparison of the Mutation Spectrum of Hypervariable Segment 1 for Phylogeographical Groups of Human Mitochondrial DNA

The nucleotide sequence variation of hypervariable segment 1 (HVS1) was analyzed for mtDNAs of 88 phylogeographical clusters characteristic of African, West Eurasian, or East Eurasian populations. A significant difference in the distribution of mutations was revealed for the mitochondrial gene pools of the regional human populations. The HVS1 positions were identified whose instability is explained by strand displacement during mtDNA replication. Strand displacement was assumed to be a major mechanism of context-dependent mutagenesis associated with the regional differentiation of human populations.