Common Region Of Allelic Loss Research Articles

Allelic loss of 9p21.3 is a prognostic factor in 1p/19q codeleted anaplastic gliomas.

We aimed to study the potential clinical relevance of 9p allelic loss, with or without copy number variation, in 1p/19q codeleted anaplastic oligodendroglial tumors (AOTs). This study enrolled 216 patients with 1p/19q codeleted AOT. The prognostic value of 9p allelic loss was investigated using a French nation-wide prospective registry, POLA (prise en charge des tumeurs oligodendrogliales anaplasiques) and high-density single nucleotide polymorphism arrays. We validated our results using the Repository of Molecular Brain Neoplasia Data (REMBRANDT) dataset. The minimal common region of allelic loss in chromosome arm 9p was 9p21.3. Allelic loss of 9p21.3, detected in 41.7% of tumors, was associated with shorter progression-free and overall survival rates in univariate (p = 0.008 and p < 0.001, respectively) and multivariate analyses (p = 0.009 and p = 0.009, respectively). This finding was validated in the REMBRANDT dataset in univariate and multivariate analysis (p = 0.01 and p = 0.01, respectively). Our study highlights a novel potential prognostic biomarker in 1p/19q codeleted AOT. Further prospective studies are warranted to investigate our finding.

Identification of a novel candidate gene, CASC2, in a region of common allelic loss at chromosome 10q26 in human endometrial cancer.

Allelic deletions, which are suggestive for the presence of tumor suppressor genes, represent a common event in endometrial cancer (EC). Previous loss-of-heterozygosity studies for human chromosome 10q identified a candidate deletion interval at 10q25-q26, which we further narrowed to a 160-kb region at 10q26, bounded by markers D10S1236 and WIAF3299. Using a positional candidate approach, we identified three alternative transcripts of a novel human gene, CASC2 (cancer susceptibility candidate 2; formely C10orf5). One of such transcripts, CASC2a, encodes a short protein of 102 amino acids with no similarity to any other known gene product. Three (7%) CASC2a mutations were identified in tumor DNA from 44 EC patients. While c.-156G>T and c.22C>T (p.Pro8Ser) are sequence variants with unknown functional significance, c.84delA is a mutation with a truncation effect on the predicted protein (p. Asn28fsX50). Expression studies by real-time RT-PCR on several normal and tumor cells revealed that CASC2a mRNA is downregulated in cancer, suggesting that it may act as a potential tumor suppressor gene. The very low mutation rate seems to also indicate that inactivation of CASC2a might probably be due to mechanisms different from genetic alterations.

Allelic Loss of Chromosome 6q in Gastric Carcinoma

Loss of the long arm of chromosome 6 (6q) has frequently been reported in gastric carcinoma, and most gastric cancer patients have evidence of intestinal metaplasia in the stomach. However, the relationship between loss of chromosome 6q and intestinal metaplasia has not been studied. In the first part of the study, we define the critical deletion region of chromosome 6q using loss of heterozygosity technique (LOH). Seventeen microsatellite markers were used to detect loss of heterozygosity (LOH) in 37 microdissected gastric tumors. We also examined intestinal metaplasia (IM) foci of the stomach in the same cancer patient (17 cases). Losses on chromosome 6q were detected in high frequency (51%) by LOH. Two distinct regions of common allelic loss were identified: one centered on the marker D6S300 (at 6q16.1) and the second on D6S446 (at 6q27), with LOH frequency of 36% and 31.3%, respectively. The deletions fall into 2 discrete regions, suggesting the existence of at least 2 tumor suppressor genes in 6q. The losses at 6q27 were confirmed by fluorescence in situ hybridization study (FISH). In the cases with LOH in the tumor, no LOH were detected in the autologous IM areas, but losses were detected by FISH. In some cases, these genetic changes may be acquired in the transition from normal gastric mucosa to intestinal metaplasia.

Frequent allelic losses on the short arm of chromosome 1 and decreased expression of the p73 gene at 1p36.3 in squamous cell carcinoma of the oral cavity

Frequent loss of heterozygosity (LOH) on the short arm of chromosome 1 (1p) has been reported in a series of human malignancies. To investigate the possible existence of tumor suppressor locus (or loci), we examined 41 primary oral squamous cell carcinomas (OSCCs) for LOH using a panel of 15 polymorphic microsatellite markers located on 1p. LOH was observed in 30 of 41 cases (73%) that were informative for at least one of the loci analyzed. Two distinct regions of common allelic loss were identified: a distal region at D1S243 (1p36.3), and proximal region at D1S160 (1p36.1). In addition, the possible involvement of the p73, a candidate tumor suppressor gene located on 1p36.3, was also evaluated. Polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis revealed no mutation of the gene in all samples analyzed (n=41). On the other hand, semi-quantitative reverse transcription-PCR (RT-PCR) demonstrated that 25% of primary tumors (n=20) had absent or reduced mRNA expression of the p73 gene. All cases showing down-regulation of the p73 gene were clinically classified as stage IV, but we could not detect any LOH at the gene locus in the same samples. Furthermore, re-expression of the p73 gene mRNA was induced in OSCC-derived cell lines showing down-regulation of the gene expression after treatment with 5-aza-2'-deoxycytidine, a DNA demethylating agent. These findings suggest that there may be at least two distinct tumor suppressor genes inactivated by allelic deletion on 1p31.1 and 1p31.3, respectively. In addition, the p73 gene could be inactivated by the methylation-dependent silencing of this gene, and associated with the tumor progression of human OSCC.

Deletion mapping of 14q32 in human neuroblastoma defines an 1,100‐kb region of common allelic loss

Deletion mapping of 14q32 in human neuroblastoma defines an 1,100‐kb region of common allelic loss

Detailed deletion mapping of chromosome band 14q32 in human neuroblastoma defines a 1.1-Mb region of common allelic loss.

Neuroblastoma (NB) is a well-known malignant disease in infants, but its molecular mechanisms have not yet been fully elucidated. To investigate the genetic contribution of abnormalities on the long arm of chromosome 14 (14q) in NB, we analysed loss of heterozygosity (LOH) in 54 primary NB samples using 12 microsatellite markers on 14q32. Seventeen (31%) of 54 tumours showed LOH at one or more of the markers analysed, and the smallest common region of allelic loss was identified between D14S62 and D14S987. This region was estimated to be 1-cM long from the linkage map. Fluorescence in situ hybridization also confirmed the loss. There was no statistical correlation between LOH and any clinicopathologic features, including age, stage, amplification of MYCN and ploidy. We further constructed a contig spanning the lost region using bacterial artificial chromosome and estimated this region to be approximately 1.1-Mb by pulsed-field gel electrophoresis. Our results will contribute to cloning and characterizing the putative tumour-associated gene(s) in 14q32 in NB. © 2000 Cancer Research Campaign

Deletion mapping of 14q32 in human neuroblastoma defines an 1,100-kb region of common allelic loss.

We analyzed loss of heterozygosity (LOH) in 54 primary neuroblastomas (NBs) using 12 microsatellite markers on 14q32, and found that 31% (17/54) NBs showed LOH. The smallest region of overlap (SRO) was identified between D14S62 and D14S987. There was no statistical correlation between LOH and any clinicopathologic features, including age, stage, amplification of MYCN, and ploidy. A sequence-ready bacterial artificial chromosome (BAC) contig was constructed, and the minimum tiling path of six BACs covered the SRO; the physical length of this region was no larger than 1,000 kb. Our findings support the existence of a putative tumor-related gene on 14q32 for the tumorigenesis of NB.

Allelic Loss at Chromosome Band 6q14 Correlates with Favorable Prognosis in Hepatocellular Carcinoma

Cytogenetic and molecular studies have frequently shown chromosome 6q deletions in non-Hodgkin lymphoma and several human cancers. There have been few studies concerning chromosome 6q deletion in hepatocellular carcinoma (HCC), and most of these studies have focused on region 6q26 ∼27. We previously described frequent allelic loss at 6q14 in HCC. As a step toward narrowing the scope of search for tumor suppressor genes, we used a series of yeast artificial chromosome clones that map to the long arm of chromosome 6 (6q14–6q22) by fluorescence in situ hybridization (FISH) to define the minimal common region of allelic loss in 25 cases of HCC. Altogether, 12 tumors had allelic loss on 6q (48%). Eleven of the 12 tumors had polysomy of chromosome 6 with evident intratumor cytogenetic heterogeneity. The minimal common region of allelic loss lies within a 2-cM region at 6q14, flanked by D6S458 ( 849_d_8) and D6S275 (911_a_3). Clinicopathologic correlation between the 12 patients with allelic loss at 6q and the 13 patients without allelic loss showed no significant differences in any basic characteristics except survival. Patients with allelic loss at 6q had a much longer median survival time than those without allelic loss (50 months vs. 11 months, P = 0.0019). Only 5 of the 25 HCC patients were still alive at the time of this study, and all of them had allelic loss at 6q, which is also statistically significant (P = 0.037, alive vs. dead). The association of allelic loss at 6q with polysomy implies that this may be a progression-associated event in HCC. The correlation of allelic loss at 6q with long survival suggests a complex mechanism of tumorigenesis in HCC and is worthy of further investigation.

Identification of two distinct deleted regions on the short arm of chromosome 1 and rare mutation of the CDKN2C gene from 1p32 in oligodendroglial tumors.

Oligodendroglial tumors frequently show allelic losses on the short arm of chromosome 1. To narrow down the putative tumor suppressor gene site(s) on 1p, we have investigated 35 oligodendrogliomas and 10 mixed gliomas (oligoastrocytomas) for loss of heterozygosity (LOH) at 21 highly polymorphic loci on chromosome 1 (19 loci on 1p and 2 loci on 1q). LOH at loci on 1p was found in 30 of the 45 tumors (67%). Two distinct regions of common allelic loss were identified: a distal region between D1S76 and D1S253 at 1p36.3, and a proximal region between D1S482 and D1S2743 at 1p34-p35. We also analyzed our tumor series for genetic alterations and expression of the cyclin dependent kinase inhibitor gene CDKN2C (p18INK4c) from 1p32. We found 1 recurrent anaplastic oligodendroglioma that carried a somatic CDKN2C mutation at codon 113 (GAA ==> TAA: Glu ==> Stop). The remaining 44 tumors of our series showed neither coding sequence mutations nor homozygous deletions of CDKN2C. Investigation of 35 tumors by differential reverse transcription-PCR revealed expression of CDKN2C transcripts in all instances. Our data thus provide evidence for more than a single oligodendroglioma-associated tumor suppressor gene on 1p and implicate CDKN2C as a candidate tumor suppressor gene altered in a low fraction of oligodendroglial tumors.

Identification of a 5-cM region of common allelic loss on 8p12-p21 in human breast cancer and genomic analysis of the hEXT1L/EXTR1/EXTL3 gene in this locus.

The short arm of chromosome 8 is frequently lost in many human carcinomas including breast cancer, suggesting the presence of a tumor suppressor gene(s) in this region. We identified a gene termed hEXT1L/EXTR1/EXTL3 (hEXT1L hereinafter) that was mapped to chromosome bands 8p12-p21 where frequent LOHs of this region was reported in breast cancer. The existence of the third breast cancer susceptibility gene was also suggested in this region by linkage analysis. We further performed LOH analysis in 8p12-p21 in 34 breast cancers and identified a 5-cM region of common allelic loss that overlapped with the locus for positive lod score in familial breast cancer. We further analyzed genomic alterations of hEXT1L in tumors in which frequent LOHs of 8p were reported. A total of 327 cancers (313 primary tumors and 14 cancer cell lines) including 22 primary breast cancers were analyzed, but none of the tumors had somatic mutations: only one thyroid cancer patient without any family history of cancer had a 9-bp insertion in the constitutional DNA. These results suggest that mutations of hEXT1L do not play a major role in the development of sporadic cancers including breast cancer, and that other tumor suppressor gene(s) exists in the 5-cM region identified in this study.

Identification of a 100-kb region of common allelic loss on chromosome bands 10q25-q26 in human endometrial cancer

Identification of a 100-kb region of common allelic loss on chromosome bands 10q25-q26 in human endometrial cancer

Allelic loss mapping and physical delineation of a region harboring a putative thymic lymphoma suppressor gene on mouse chromosome 12.

Our previous allelic loss analysis of gamma-ray induced thymic lymphomas in F1 hybrid and backcross mice between BALB/c and MSM strains mapped the Tlsr4 region exhibiting a high frequency of allelic loss (62%) to a 2.9 cM interval between the markers D12Mit53 and D12Mit279 on mouse chromosome 12. To narrow further the interval harboring a putative tumor suppressor gene, a high-density scan has been carried out for informative 361 thymic lymphomas. Construction of a physical map of Tlsr4 with 3 YAC and 15 BAC clones and isolation of YAC- and BAC-derived polymorphic probes lead to fine allelic loss mapping. Three successive polymorphic sites within one BAC exhibit the retention of both alleles in seven, one and four lymphomas, suggesting that a common region of allelic loss for Tlsr4 exists within the BAC region. Pulsed-field gel electrophoresis of NotI digests of this and other clones determines that the commonly lost region is a 35 kb interval with a NotI site. NotI sites are frequently associated with coding regions, and our preliminary sequencing has identified ESTs in the region. Thus, the present study facilitates the identification of genes in the Tlsr4 region that would lead to isolation of a novel tumor suppressor gene.

Localization of common deletion regions on 1p and 19q in human gliomas and their association with histological subtype.

Allelic alterations of chromosomes 1 and 19 are frequent events in human diffuse gliomas and have recently proven to be strong predictors of chemotherapeutic response and prolonged survival in oligodendrogliomas (Cairncross et al., 1998; Smith et al., submitted). Using 115 human diffuse gliomas, we localized regions of common allelic loss on chromosomes 1 and 19 and assessed the association of these deletion intervals with glioma histological subtypes. Further, we evaluated the capacity of multiple modalities to detect these alterations, including loss of heterozygosity (LOH), fluorescence in situ hybridization (FISH), and comparative genomic hybridization (CGH). The correlation coefficients for detection of 1p and 19q alterations, respectively, between modalities were: 0.98 and 0.87 for LOH and FISH, 0.79 and 0.60 for LOH and CGH, and 0.79 and 0.53 for FISH and CGH. Minimal deletion regions were defined on 19q13.3 (D19S412-D19S596) and 1p (D1S468-D1S1612). Loss of the 1p36 region was found in 18% of astrocytomas (10/55) and in 73% (24/33) of oligodendrogliomas (P < 0.0001), and loss of the 19q13.3 region was found in 38% (21/55) of astrocytomas and 73% (24/33) of oligodendrogliomas (P = 0.0017). Loss of both regions was found in 11% (6/55) of astrocytomas and in 64% (21/33) of oligodendrogliomas (P < 0.0001). All gliomas with LOH on either 1p or 19q demonstrated loss of the corresponding FISH probe, 1p36 or 19q13.3, suggesting not only locations of putative tumor suppressor genes, but also a simple assay for assessment of 1p and 19q alterations as diagnostic and prognostic markers.

Identification of three commonly deleted regions on chromosome arm 6q in human pancreatic cancer

Pancreatic cancer has one of the poorest prognoses among malignant diseases. To understand its molecular mechanisms, we studied allelic losses on the long arm of chromosome 6. Using 55 paired DNAs of tumors and their corresponding normal tissues and 30 microsatellite markers that spanned the entire 6q chromosome arm, we found three distinct regions of common allelic loss: region A, a less than 500-kb region bordered by D6S449 and D6S283 on 6q21 with a loss of heterozygosity (LOH) frequency of 69% (38/55); region B, a 7-cM region bordered by D6S292 and D6S308 on 6q23-q24 with a LOH frequency of 60% (33/55); and region C, a 13-cM region bordered by D6S305 and D6S264 with a LOH frequency of 51% (28/55). We further focused on region A and constructed a physical map using yeast artificial chromosome (YAC) clones, their derived cosmid clones, and bacterial artificial chromosome (BAC) clones. Region A was completely covered by three overlapping BAC clones. Our results in the present study should shed light on the cloning and characterization of tumor suppressor genes in pancreatic carcinogenesis.

The FEZ1 gene at chromosome 8p22 encodes a leucine-zipper protein, and its expression is altered in multiple human tumors.

Alterations of human chromosome 8p occur frequently in many tumors. We identified a 1.5-Mb common region of allelic loss on 8p22 by allelotype analysis. cDNA selection allowed isolation of several genes, including FEZ1. The predicted Fez1 protein contained a leucine-zipper region with similarity to the DNA-binding domain of the cAMP-responsive activating-transcription factor 5. RNA blot analysis revealed that FEZ1 gene expression was undetectable in more than 60% of epithelial tumors. Mutations were found in primary esophageal cancers and in a prostate cancer cell line. Transcript analysis from several FEZ1-expressing tumors revealed truncated mRNAs, including a frameshift. Alteration and inactivation of the FEZ1 gene may play a role in various human tumors.

Identification of two distinct deletion targets at 11q23 in cutaneous malignant melanoma.

Karyotypic and molecular data indicate that genetic alterations of the long arm of chromosome 11 (11q) are involved in the pathogenesis of malignant melanoma as well as of other malignancies. We have shown previously, by analysis of loss of heterozygosity (LOH), that a tumor-suppressor gene playing an important role in malignant melanoma is likely to be located within a 51-cM region at 11q23. Its loss appeared to be a late event in tumor progression and an indicator of a less favorable clinical outcome. To further test this hypothesis on a larger set of tumors and to refine the region(s) of common allelic loss, we analyzed 21 polymorphic microsatellite repeats on 11q. A PCR-based assay for LOH was used to study normal and tumor tissues from 53 individuals with primary cutaneous malignant melanoma or metastatic disease. Our findings indicate that in cutaneous malignant melanoma there are at least 2 distinct regions of common allelic loss on 11q, one of them centered around marker APOC3 at 11q23.1-q23.2 delineated by markers D11S1347 and D11S4142 and spanning approximately 5 Mb and a second 3-Mb region around marker D11S925 at 11q23.3 delineated by markers D11S528 and D11S1345. Both regions have been described as deletion targets or as being included within larger allelic deletions detected in several other common tumor types. Thus, these 2 putative melanoma-suppressor loci are likely to harbor tumor-suppressor genes relevant to tumorigenesis of melanoma and a number of other common human malignancies.

Detailed deletion mapping on chromosome 1p32-p36 in human colorectal cancer: identification of three distinct regions of common allelic loss.

Recent studies have suggested the existence of one or several tumor-suppressor genes on chromosome arm 1p in colorectal tumors. To determine the localization of the putative tumor suppressor genes, we performed LOH analysis in 1p in colorectal tumors. A total of 48 paired normal and tumor DNAs of 46 colorectal tumor patients and 21 microsatellite markers on 1p32.1-p36.3 were used for PCR-LOH analysis. Three commonly deleted regions were found: i) 1p36.3 (10-cm); ii) 1p35.1-p36.3 (2-cm); and iii) 1p34.2-p35 (1-cm). These regions overlapped with those reported in several types of tumor. No significant associations were found between LOH and clinicopathologic features. The regions identified in the present study could harbor tumor suppressor genes that would also be associated with several types of human cancer.

Identification of a 100-kb region of common allelic loss on chromosome bands 10q25-q26 in human endometrial cancer.

In human endometrial cancer, we have previously identified a 790-kb region of common allelic loss in chromosome bands 10q25-q26, flanked by D10S587 and D10S1723. We constructed a contig covering the entire deleted region using YACs, PACs, and BACs. Five overlapping cosmid clones derived from YAC clones completely covered the entire deleted region: its size was estimated to be no larger than 200 kb. We further performed two-color fluorescence in situ hybridization (FISH) analysis to confirm the deletion and narrowed down the deleted region to 100 kb or less; it was covered by three overlapping cosmid clones that were included in one BAC clone. Restriction endonuclease mapping identified a region in which NotI, SalI, SmaI, and Xhol were clustered, suggesting the possible existence of a CpG island.

A common region of allelic loss on chromosome region 3p25.3-26.3 in nasopharyngeal carcinoma.

Loss of heterozygosity (LOH) of chromosome arm 3p has been commonly observed in carcinomas of various tissues, including those of nasopharyngeal carcinoma (NPC). To determine the frequency and extent of allelic loss in NPC, we investigated 16 loci on chromosome bands 3p21-26 in 24 tumor tissues by microsatellite analysis. LOH on 3p21-26 was found in 16 of 24 (66.7%) tumors. The highest frequency of allelic loss was found in two adjacent loci, D3S1620 (11/22, 50%) and D3S1560 (9/18, 50%). Eight cases showed LOH in one contiguous region and 5 cases in more than one region. Samples 1, 3, 4, 7, 8, 10, 16, 17, 18, 19, and 22 had a contiguous stretch of allelic loss between D3S1297 and D3S1597. The smallest common LOH/deletion region seems likely to lie between D3S1297 (3p26.3-26.2) and D3S1560 (3p25.3). The allelic loss map defined here will facilitate finer mapping of putative tumor suppressor gene loci and positional cloning of such genes, which may play a role in carcinogenesis of NPC.

The H-cadherin (CDH13) gene is inactivated in human lung cancer.

We have previously reported frequent allelic loss in chromosome bands 16q24.1-q24.2 in human lung cancer. Since the H-cadherin (CDH13) gene has been isolated and mapped to this common region of allelic loss, we have investigated this gene in human lung cancer. The reverse transcription/polymerase chain reaction technique has revealed the loss of expression in four (57%) of seven lung cancer cell lines. To study the CDH13 gene further, we have analyzed deletions, genetic alterations, and methylation status at the 5' region of this gene. Three (75%) of four cell lines that have lost expression show a deletion of the CDH13 locus accompanied by hypermethylation of the remaining allele. Moreover, hypermethylation has been observed in nine (45%) of 20 primary lung cancers. These results suggest that a combination of deletion and hypermethylation causes inactivation of the CDH13 gene in a considerable number of human lung cancers.