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LOH Analysis (loh + analysis)
Selected AbstractsNarrowing of the regions of allelic losses of chromosome 1p36 in meningioma tissues by an improved SSCP analysisINTERNATIONAL JOURNAL OF CANCER, Issue 8 2008Yanlei Guan Abstract Mapping loss of heterozygosity (LOH) regions in the genomes of tumor tissues is a practical approach for identifying genes whose loss is related to tumorigenesis. Conventional LOH analyses using microsatellite or single nucleotide polymorphism (SNP) markers require the simultaneous examination of tumor- and matched normal-DNA. Here, we improved the previously developed SNP-based LOH assay using single strand conformation polymorphism (SSCP) analysis, so that LOH in tumor samples heavily contaminated with normal DNA can now be precisely estimated, even when matched normal DNA is not available. We demonstrate the reliability of the improved SSCP-based LOH detection method, called the LOH estimation by quantitative SSCP analysis using averaged control (LOQUS-AC), by comparing the results with those of the previous "LOH estimated by quantitative SSCP assay" (LOQUS) method. Using the LOQUS-AC assay, LOH was detected at a high consistency (98.1%) with the previous LOQUS method. We then applied this new method to characterize LOH profiles in 130 meningiomas, using 68 SNPs (i.e., a mean inter-SNP interval of 441 kbp) that are evenly distributed throughout chromosome 1p36. Benign, atypical and anaplastic meningiomas exhibited 1p36 LOH at frequencies of 48.39, 84.62 and 100.00%, respectively, using LOQUS-AC. Subsequently, we detected a candidate common LOH region on 1p36.11 that might harbor tumor suppressor genes related to malignant progression of meningioma. © 2007 Wiley-Liss, Inc. [source] 3,-azido-3,-deoxythymidine induces deletions in L5178Y mouse lymphoma cells,,ENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 3-4 2007Jianyong Wang Abstract 3,-Azido-3,-deoxythymidine (AZT), a nucleoside analogue used for the treatment of acquired immunodeficiency syndrome (AIDS), induced a significant dose-related increase in the thymidine kinase (Tk) mutant frequency (MF) in L5178Y/Tk+/, 3.7.2C mouse lymphoma cells. Treatment with 1 mg/ml (3,742 ,M) AZT for 24 hr resulted in a MF of 407 × 10,6 compared to a control MF of 84 × 10,6. The MFs of the large and small colony mutants resulting from AZT exposure were 142 × 10,6 and 265 × 10,6, respectively. One hundred and fifty mutants from the 1 mg/ml (3,742 ,M) AZT-treated culture and sixty-nine mutants from independent untreated cultures were isolated and analyzed. LOH analysis using a heteromorphic microsatellite locus located in the Tk gene was performed to determine the presence or absence of the Tk+ allele. Eight other microsatellite markers spanning the entire mouse chromosome 11 also were examined for heterozygosity to determine the extent of LOH. In addition, Tk gene dosage analysis was conducted using Real-Time PCR in those mutants showing LOH at the Tk locus. The presence of only one Tk allele based on Real-Time PCR indicated that the mutant resulted from deletion while the presence of two alleles was consistent with a recombination event. More mutants from the AZT-treated culture showed Tk LOH than did independent mutants from the untreated cultures (91% vs. 64%) and the induced mutants also showed distinct chromosome 11 LOH patterns. The mutation spectrum of mutants from AZT-treated cells was also significantly different from that of spontaneous mutants. More deletions and fewer intragenic mutations were observed in the mutants from the AZT-treated culture than independent mutants from the untreated control. Our data indicate that AZT primarily induced LOH mutations in L5178Y mouse lymphoma cells and a large number of LOH mutations resulted from deletions. Environ. Mol. Mutagen., 2007. Published 2007 Wiley-Liss, Inc. [source] Genome-wide amplification and allelotyping of sporadic pituitary adenomas identify novel regions of genetic lossGENES, CHROMOSOMES AND CANCER, Issue 3 2003D. J. Simpson Through the use of a candidate gene approach, several previous studies have identified loss of heterozygosity (LOH) at putative tumor-suppressor gene (TSG) loci in sporadic pituitary tumors. This study reports a genome-wide allelotyping by use of 122 microsatellite markers in a large cohort of tumors, consisting of somatotrophinomas and non-functioning adenomas. Samples were first subject to prior whole genome amplification by primer extension pre-amplification (PEP) to circumvent limitations imposed by insufficient DNA for whole-genome analysis with this number of microsatellite markers. The overall mean frequency of loss in invasive tumors was significantly higher than that in their non-invasive counterparts (7 vs. 3% somatotrophinomas; 6 vs. 3% non-functioning adenomas, respectively). Analysis of the mean frequency of LOH, across all markers to individual chromosomal arms, identified 13 chromosomal arms in somatotrophinomas and 10 in non-functioning tumors, with LOH greater than the 99% upper confidence interval calculated for the rate of overall random allelic loss. In the majority of cases, these losses were more frequent in invasive tumors than in their non-invasive counterparts, suggesting these to be markers of tumor progression. Other regions showed similar frequencies of LOH in both invasive and non-invasive tumors, implying these to be early changes in pituitary tumorigenesis. This genome-wide study also revealed chromosomal regions where losses were frequently associated with an individual marker, for example, chromosome arm 1q (LOH > 30%). In some cases, these losses were subtype-specific and were found at a higher frequency in invasive tumors than in their non-invasive counterparts. Identification of these regions of loss provides the first preliminary evidence for the location of novel putative TSGs involved in pituitary tumorigenesis that are, in some cases, subtype-specific. This investigation provides an unbiased estimate of global aberrations in sporadic pituitary tumors as assessed by LOH analysis. The identification of multiple "hotspots" throughout the genome may be a reflection of an unstable chromatin structure that is susceptible to a deletion or epigenetic-mediated gene-silencing events. © 2003 Wiley-Liss, Inc. [source] Loss of heterozygosity analysis: Practically and conceptually flawed?GENES, CHROMOSOMES AND CANCER, Issue 4 2002Ian P.M. Tomlinson The Knudson "two-hit" hypothesis has provided the rationale for studies that aim to identify tumor-suppressor genes by mapping regions of allelic loss (loss of heterozygosity, LOH). Although LOH has been found in practically all types of tumors, very few such projects have been successful in identifying their tumor-suppressor targets. The prime explanation for this failure is probably that researchers have, in general, been too credulous about the two-hit hypothesis, and too willing to ignore factors such as intratumor heterogeneity, contamination by normal cells, karyotypic complexity, homozygous deletions, gene dosage changes, and polymerase chain reaction artifacts. We suggest ways of minimizing these problems. Unfortunately, there is no guarantee that existing or newer methods, such as genomic microarrays and in situ single-nucleotide polymorphism analysis, will solve the difficulties of LOH analysis. The future prospects for LOH studies are, as ever, uncertain. © 2002 Wiley-Liss, Inc. [source] Chromosome 9 deletions are more frequent than FGFR3 mutations in flat urothelial hyperplasias of the bladderINTERNATIONAL JOURNAL OF CANCER, Issue 5 2006Johanna M.M. van Oers Abstract Flat urothelial hyperplasias (FUHs) in patients with papillary bladder tumours frequently show deletions of chromosome 9, suggesting that FUH could be the first neoplastic step in the development of papillary bladder cancer. FGFR3 mutations are frequent in non-invasive papillary tumours with low risk of progression. Our aim was to investigate the frequency of FGFR3 mutations and deletions of chromosomes 9p/q and 8p/q in FUH. Thirty FUH and 9 simultaneous or consecutive tumours were detected by 5-ALA-based photodynamic cystoscopy. DNA was isolated from frozen sections and whole genome amplification was done by I-PEP-PCR, followed by LOH analysis on chromosomes 8p/q and 9p/q. FGFR3 mutations were detected by SNaPshot analysis. LOH analysis on FUH revealed deletions at 9p/q (11/30, 37%) and 8p/q (3/30, 10%). FGFR3 mutations were found in 7/30 FUH (23%). Only 2 FUH showed an FGFR3 mutation without deletions of chromosome 9. In contrast, 6 FUH revealed chromosome 9 deletions but wild type FGFR3 (p = 0.03). These results suggest that chromosome 9 deletions are the earliest genetic alterations in bladder cancer. The detection of FGFR3 mutations in FUH further supports the role of this lesion as precursor of papillary bladder cancer. © 2006 Wiley-Liss, Inc. [source] | ||||||||||