Deletion mapping of the short arm of chromosome 8 in non-small cell lung carcinoma

Frequent losses of heterozygosity observed at several chromosomal loci in primary lung cancers have indicated the existence of several tumor suppressor genes associated with this type of cancer. We have examined loss of heterozygosity on chromosomal arm 8p in 49 cases of non-small cell lung carcinoma, using 14 restriction fragment length polymorphism markers. Of 42 cases informative with at least one marker, 21 showed allelic loss, including 15 of 32 adenocarcinomas and 5 of 9 squamous cell carcinomas. The frequency of allelic loss on 8p was similar at all clinical stages. Deletion mapping defined a single common region of deletion in these tumors within an 8 cM interval at 8p21.3–p22 flanked by the loci defined by cMSR-32 and cC18–245. © 1993 Wiley-Liss, Inc.     

Deletion mapping of the short arm of chromosome 3 in Merkel cell carcinoma

Little is known about the biology of Merkel cell carcinoma (MCC), also called small cell carcinoma of the skin. MCC has similarities with small cell lung cancer (SCLC): both are neuroendocrine malignancies with early metastasis to distant sites and a poor prognosis. Small cell lung cancer biopsies are known to have frequent losses on chromosome 3 in the region 3p21, yet MCCs have not been reported to have 3p deletions by karyotypic analysis. Considering the similarities between SCLC and MCC, we investigated 26 MCC tumours for loss of heterozygosity (LOH) on 3p. First, RFLP analysis was performed using PCR with nine primer sets from six loci. Second, 25 tumours were examined by microsatellite analysis for 3p markers D3S1289 and D3S1285 and SST on 3q. All 26 tumours were informative at one or more loci; of these, 18 (69%) demonstrated LOH for at least one marker on the short arm. For all informative loci the frequency of LOH was greater than 30% (range 33–75%). In a cell line derived from one tumour, it was possible to demonstrate rearrangement of chromosome 3 by in situ hybridisation. No LOH was seen in 15 informative cases for the 3q locus SST. A region 3p13-p21.1, centered on the marker D3S2, was deleted in all tumours demonstrating LOH, with a secondary deletion involving D3S30 detected in some tumours at 3p13. Our results indicate that LOH on 3p is a common occurrence in MCC; however, three tumours for which DNA was also available from a corresponding cell line suggest there may be a subset of MCC whose genesis is independent of deletions of 3p. Genes Chromosom Cancer 15:102–107 (1996) © 1996 Wiley-Liss, Inc.     

Deletion mapping of the short arm of chromosome 3 in human malignant mesothelioma

Previous cytogenetic investigations have revealed frequent deletions and ocher unbalanced structural rearrangements of 3p in human malignant mesothelioma. We have performed a restriction fragment length polymorphism analysis by using the polymerase chain reaction and primer sets for seven DNA markers to examine loss of heterozygosity (LOH) from 3p in 25 malignant mesotheliomas. Among 24 cases informative at one or more 3p loci, 15 (62.5%) exhibited LOH with at least one marker. Deletion mapping in these tumors indicates that the common region of chromosomal loss resides within band 3p21, in the vicinity of the D3F15S2 locus. These results suggest that allelic loss from 3p21 is a frequent Occurrence in malignant mesothelioma and that one or more putative tumor suppressor genes at this site contribute to the pathogenesis of this malignancy.Genes Chrom Cancer 9:76-80 (1994). © 1994 Wiley-Liss, Inc.     

Deletion mapping of chromosomes 8, 10, and 16 in human prostatic carcinoma

In an allelotyping study prostatic carcinoma, we found the highest frequency of allelic deletions on chromosomes 8, 10, 16, and 18. In all cases with allelic deletions, at least one of the chromosomes 8, 10, and 16 were involved. A detailed deletion mapping of these chromosomes in 18 cases was carried out with probes that detect restriction fragment length polymorphisms (RFLP) on chromosomes 8 (6 probes), 10 (11 probes), and 16 (9 probes). The highest frequency of allelic deletions were found on 8p (65%), where the minimally deleted region was between the PLAT locus and pter. The long arm of chromosome 16 had allelic deletions in 56% of informative cases, with three different break points, the most distal being located between D16S4 and D16S7. Chromosome 10 exhibited a complex deletion pattern with terminal deletions of the p or the q arm (2 cases each), a deletion pattern that could be interpreted as nonreciprocal translocations of the q arm (2 cases), or allelic losses on all informative loci, suggesting monosomy (2 cases). Our data suggest that tumor suppressor genes involved in the oncogenesis of prostatic carcinoma may be localized between 8 pter and the PLAT locus and that additional/alternative tumor suppressor genes may be localized on chromosome 10 and on the long arm of chromosome 16 distal to the D16S4 locus.     

Deletion mapping of the long arm of chromosome 10 in glioblastoma multiforme

Cytogenetic and restriction fragment length polymorphism (RFLP) studies have shown that loss of one entire copy of chromosome 10 is a common genetic event in glioblastoma multiforme, the most malignant glial brain tumor in humans. In a search for submicroscopic deletions, we carried out an RFLP analysis using markers that had been mapped accurately on chromosome 10 by genetic linkage analysis. We studied 30 patients of whom 15 had loss of heterozygosity (LOH) at one or more loci. In seven cases, LOH was found at every informative locus, whereas in two cases extensive deletions were observed involving both the short and long arms. In six other patients, LOH was confined to a portion of the long arm. The smallest region of overlap among these latter six deletions was flanked by markers D10S12 proximally and D10S6 distally, a 33.4 centimorgan region that maps physically near the telomere (q25.1-qter). This region will serve as an important target for future mapping experiments designed to identify a tumor suppressor gene implicated in this lethal form of human cancer. Genes Chrom Cancer 7:173–177 (1993). © 1993 Wiley-Liss, Inc.     

Deletion mapping of 18q in conventional renal cell carcinoma

Loss of heterozygosity (LOH) is frequently associated with the inactivation of tumor suppressor genes. 18q LOH has been frequently reported in colorectal cancer and lung cancer; however, allelic loss on 18q has not been investigated in renal cell carcinoma (RCC). We evaluated LOH on 18q using nine microsatellite markers in 126 with conventional RCC (cRCC). LOH was observed in more than one 18q microsatellite locus in 24 cRCC (19%). We found the highest frequency of LOH (13.5%) at 18q21.3, where the DCC gene is located. We also assessed the relationship between LOH frequency and patient clinical parameters. Patients with a family history of cancer had a significantly higher frequency of 18q LOH than those without such a history (P=0.0017). No associations were found with other parameters, including gender, tumor grade, tumor stage, smoking status, and body mass index. The results suggest that inactivation of tumor suppressor genes at 18q21.3, including DCC and SMAD4 as candidates, may be involved in the tumorigenesis of some conventional RCCs.     

Deletion mapping of chromosome region 12q13-24 in colorectal cancer

Colorectal cancer is one of the most common cancers in the world. Colorectal cancer develops after a long and multistep process of carcinogenesis. Inactivation of tumor suppressor genes is among the most important steps in development of colorectal cancer. Analysis of loss of heterozygosity (LOH) is an effective method to determine the localization of tumor suppressor genes. In this study, we used five microsatellite markers to analyze the region 12q13-24 among 47 patients with colorectal cancer. The frequency of LOH and the clinicopathological data were compared using logistic regression and a chi-square test. In 34 of 47 tumor tissues (72%), LOH was detected at least in one marker. The highest LOH frequency was 34%, on the D12S129 locus; the lowest frequency was 23%, on the D12S78 locus. Loss of heterozygosity was detected as 32% on D12S83, 30% on D12S346, and 26% on D12S1660. No statistically significant correlation was found between the frequency of LOH and clinicopathological features (P > 0.05). Chromosome region 12q13-24 contains several known genes that may be candidate tumor suppressor genes, including RASAL1, ITGA7, STAB2, GLIPR1, and SLC5A8. Although the exact roles of these genes in colorectal cancer formation remain to be clarified, the present data point to a tumor suppressor role.     

Deletion of chromosome 21 disturbs human brain morphogenesis.

PURPOSE: Humans with small deletions of chromosome 21 provide important models for understanding the role of dosage-sensitive genes in brain morphogenesis. To identify chromosome 21 genes responsible for defects of the central nervous system, we determined the deleted regions and brain malformations in three unrelated individuals with overlapping partial deletions of chromosome 21. METHODS: Fluorescent in situ hybridization and magnetic resonance imaging were used to define the chromosomal structure and structural brain abnormalities present in these three individuals. RESULTS: The regions of chromosome 21 found to be deleted in these individuals were as follows: case 1: KCNJ6 to the telomere; case 2: ITSN1 to the telomere; and case 3: ITSN1 to PCNT2. The abnormalities of brain structure shared by all included microcephaly, pachygyria, polymicrogyria, colpocephaly, hypoplastic corpus callosum and white matter, hypoplastic cerebellum, and enlarged ventricular system. The clinical features in common included mental retardation, microcephaly, facial dysmorphism, and epilepsy (severe in one patient). CONCLUSION: From analyses of the molecular, cytogenetic, and neuroimaging data from these three individuals, combined with those from previously reported cases, we infer that deletion of an 8.4-Mb region in chromosome band 21q22.2-22.3 (KCNJ6-COL6A2) is associated with cortical dysplasia. We propose that one or more dosage-sensitive genes in this region contributes to cortical development and that deletion of 21q22.2-22.3 should be considered in the diagnosis of mentally retarded patients with facial dysmorphism and cerebral dysplasia.     

Molecular mapping of deletion sites in the short arm of chromosome 3 in human lung cancer

We used 10 restriction fragment length polymorphism (RFLP) probes spanning the length of the short arm of chromosome 3 (3p) to map deletion sites in human lung cancer. Two approaches were used. 1) When a patient's tumor and normal tissue were available, loci with allelic heterozygosity in the normal tissue were tested for loss of alleles at 3p. 2) When the corresponding normal tissue was not available, the frequency of heterozygosity at each locus in a panel of tumors was compared to the corresponding published frequencies in nontumor tissue of healthy individuals or patients with lung cancer. In 14 small cell lung carcinomas (SCLC) with normal DNA for comparison, allele loss was found at all heterozygous loci, with one exception at a locus near the 3p centromere (D3S4). In the total of 53 SCLCs, which included tumors without paired normal tissue, frequency of heterozygosity was significantly reduced in all 10 3p loci. Three loci, DNF 15S2, RAF1, and D3S18, were homozygous in all tumors in the SCLC panel. These loci, which are in regions 3p21 and 3p25, may thus be involved in the origin or evolution of SCLC. We also investigated 24 non-SCLC tumors. In this panel, frequency of heterozygosity was significantly reduced at seven of the 10 loci tested. Comparison of the results shows that the pattern of allele loss on 3p is different in SCLC and non-SCLC, suggesting a difference in pathogenesis at the genetic level.     

The origin of human chromosome 2 analyzed by comparative chromosome mapping with a DNA microlibrary

Fluorescencein situ hybridization (FISH) of microlibraries established from distinct chromosome subregions can test the evolutionary conservation of chromosome bands as well as chromosomal rearrangements that occurred during primate evolution and will help to clarify phylogenetic relationships. We used a DNA library established by microdissection and microcloning from the entire long arm of human chromosome 2 for fluorescencein situ hybridization and comparative mapping of the chromosomes of human, great apes (Pan troglodytes, Pan paniscus, Gorilla gorilla, Pongo pygmaeus) and Old World monkeys (Macaca fuscata andCercopithecus aethiops). Inversions were found in the pericentric region of the primate chromosome 2p homologs in great apes, and the hybridization pattern demonstrates the known phylogenetically derived telomere fusion in the line that leads to human chromosome 2. The hybridization of the 2q microlibrary to chromosomes of Old World monkeys gave a different pattern from that in the gorilla and the orang-utan, but a pattern similar to that of chimpanzees. This suggests convergence of chromosomal rearrangements in different phylogenetic lines.     

Deletion mapping of chromosome 13q in head and neck squamous cell carcinoma in Indian patients: correlation with prognosis of the tumour

Deletions in chromosome (chr.) 13q occur frequently in head and neck squamous cell carcinoma (HNSCC). Previous studies failed to identify common deleted regions in chr.13q, though several candidate tumour suppressor genes (TSGs) loci, e.g. BRCA2, RB1 and BRCAX have been localized in this chromosome, as well as no prognostic significance of the deletion has been reported. Thus, in the present study, deletion mapping of chr. 13q has been done in 55 primary HNSCC samples of Indian patients using 11 highly polymorphic microsatellite markers of which three were intragenic to BRCA2 gene, one intragenic to RB1 gene and another from BRCAX locus. The deletion in chr.13q was significantly associated with progression of HNSCC. High frequencies (27-39%) of loss of heterozygosity were found in 13q13.1 (BRCA2), 13q14.2 (RB1), 13q21.2-22.1 (BRCAX) and 13q31.1 regions. Deletions in the BRCA2 and RB1 regions were significantly correlated. The four highly deleted regions were associated with clinical stage and histological grades of the tumour as well as poor patient outcome. Deletion in the 13q31.1 region was only found to be associated with HPV infection. High frequencies (11-23%) of microsatellite size alteration (MA) were seen to overlap with the highly deleted regions. Forty per cent of the samples showed rare biallelic alteration whereas loss of normal copy of chromosome 13q was seen in five tumours. Thus, it seems that the putative TSGs located in the BRCAX and 13q31.1 regions as well as the BRCA2 and RB1 genes may have some cumulative effect in progression and poor prognosis of HNSCC. Significant association between deletion in BRCA2 and RB1 gene loci may indicate functional relationship between the genes in this tumour progression.     

Comparative FISH mapping of the ancestral fusion point of human chromosome 2

It is known that human chromosome 2 originated from the fusion of two ancestral primate chromosomes. This has been confirmed by chromosome banding and fluorescence in-situ hybridization (FISH) with human chromosome-2-specific DNA libraries. In this study, the order of 38 cosmid clones derived from the human chromosome region 2q12-q14 was exactly determined by high-resolution FISH in human chromosome 2 and its homologous chromosomes in chimpanzees (Pan trogrodydes, 2n=48) and cynomolgus monkeys (Macaca fascicularis, 2n=42). This region includes the telomere-to-telomere fusion point of two ancestral ape-type chromosomes. As a result of comparative mapping, human chromosome region 2q12-q14 was found to correspond to the short arms of chimpanzee chromosomes 12 and 13 and cynomolgus monkey chromosomes 9 and 15. It is noted that no difference was detected in the relative order of the cosmid clones between human and chimpanzee chromosomes. This suggests that two ancestral ape-type chromosomes fused tandemly at telomeres to form human chromosome 2, and the genomic organization of this region is thought to be considerably conserved. In the cynomolgus monkey, however, the order of clones in each homologue was inverted. In addition to cosmid mapping, two chromosome-2-specific yeast artificial chromosome (YAC) clones containing the fusion point were identified by FISH. This revised version was published online in July 2006 with corrections to the Cover Date.     

Deletion mapping on the long arm of chromosome 7 in splenic lymphoma with villous lymphocytes

Splenic lymphoma with villous lymphocytes (SLVL) is a low-grade lymphoproliferative disorder characterized by splenomegaly and circulating villous lymphocytes in the peripheral blood. It is considered to be the leukemic form of splenic marginal zone lymphoma (SMZL). The genetic basis of this lymphoma type remains unknown. Conventional cytogenetic studies have identified frequent structural abnormalities of chromosome 7, in the form of translocations, mainly unbalanced, and 7q deletions. In this current study, we undertook deletion mapping of the long arm of chromosome 7 in a series of cases with SLVL. Metaphase fluorescence in situ hybridization (FISH) was used in the first instance, followed by a study of loss of heterozygosity (LOH). The common area of deletion identified by FISH spanned from the YAC clone HSC7E1289 (mapping to 7q32.1) to in between YACs HSC7E195 and HSC7E648 (7q32-3). By application of 50 microsatellite markers mapping to the FISH-CDR and to areas of deletion reported in other studies, four distinct hotspot loci were identified, with abnormalities present in 29-55% cases. In three of them, both LOH and biallelic deletions were found. The LOH in the majority of patients was noncontiguous. The presence of a high incidence of abnormalities in the established hotspot areas and in particular the finding of biallelic deletions is indicative of the existence of genes important for the pathogenesis of SLVL in these areas.     

Optical mapping of Plasmodium falciparum chromosome 2

Detailed restriction maps of microbial genomes are a valuable resource in genome sequencing studies but are toilsome to construct by contig construction of maps derived from cloned DNA. Analysis of genomic DNA enables large stretches of the genome to be mapped and circumvents library construction and associated cloning artifacts. We used pulsed-field gel electrophoresis purified Plasmodium falciparum chromosome 2 DNA as the starting material for optical mapping, a system for making ordered restriction maps from ensembles of individual DNA molecules. DNA molecules were bound to derivatized glass surfaces, cleaved with NheI or BamHI, and imaged by digital fluorescence microscopy. Large pieces of the chromosome containing ordered DNA restriction fragments were mapped. Maps were assembled from 50 molecules producing an average contig depth of 15 molecules and high-resolution restriction maps covering the entire chromosome. Chromosome 2 was found to be 976 kb by optical mapping with NheI, and 946 kb with BamHI, which compares closely to the published size of 947 kb from large-scale sequencing. The maps were used to further verify assemblies from the plasmid library used for sequencing. Maps generated in silico from the sequence data were compared to the optical mapping data, and good correspondence was found. Such high-resolution restriction maps may become an indispensable resource for large-scale genome sequencing projects.     

Deletion of chromosome arm 1p in a merkel cell carcinoma (MCC)

A case of neuroendocrine skin carcinoma (Merkel cell carcinoma) with a deletion of the short arm of chromosome 1 (1p) as the sole chromosomal abnormality was examined. The tumor originated in the skin of the left knee of a 67-year-old man. Histopathologic study showed an undifferentiated small cell tumor which expressed neuron-specific enolase, chromogranin, and cytokeratin (CAM 5.2). Cytogenetic analysis of a lymph node metastasis from the groin showed a pseudodiploid cell population with a deletion of the short arm of chromosome 1 as the only abnormality: 46,XY,del( 1 )(p36.1). In situ hybridization with the DI22 probe specific for the terminal band of 1 p confirmed the terminal deletion. This is the first case of Merkel cell carcinoma in which only one chromosomal abnormality has been observed. Loss of the terminal portion of 1p suggests that a tumor supressor gene on 1p plays a role in the pathogenesis of Merkel cell carcinoma. Genes Chrom Cancer 9:216-220 (1994). © 1994 Wiley-Liss, Inc.     

Premature chromosome condensation in human cervical carcinoma

Premature chromosome condensation was observed in 22 of 150 cases (15%) of cervical cancers studied with G-banding. The percentage of premature chromosome condensation in the tumors ranged from 1.5% to 8.9%. Premature chromosome condensation was present in greater frequency (16 of 22 cases) in tumors showing hyperdiploidy, suggesting that this could be a major mechanism of induction of polyploidy in addition to other operative mechanisms, such as endomitosis and endoreduplication. No relationship, either with the disease stage or prognosis, was evident.     

Deletion 3p: the only chromosome loss in a primary renal cell carcinoma

The cytogenetic findings obtained by G- and C-banding in short-term culture from a primary renal cell carcinoma are reported. This tumor is characterized by loss of the short arm of chromosome #3 and t(Y;3). The aberration of chromosome #3 in renal cell carcinomas is discussed.     

Deletion mapping of chromosome 1 in early onset and late onset breast tumors--a comparative study in eastern India

Younger women exhibit more aggressive pathologic features of breast cancer (BC) compared to their older counterparts. Young age has been shown to be an independent predictor of adverse prognosis. These findings have raised the question of whether these differences are also present at the genetic level. Twenty-five early onset (age < or = 40 years) tumors including 4 bilateral tumors, and 26 late onset (>40 years) breast tumors, including 2 bilateral tumors, were examined for loss of heterozygosity (LOH) at chromosome 1 using 11 polymorphic microsatellite markers. A comparative study revealed high frequencies of LOH in chr. 1p36 (61%), 1p31.3 (40%), 1p21.3 (50%) and 1q22-23.2 (56%) in a younger group, and chr. 1p36 (46%), 1p34.2 (48%), and 1q22-23.2 (52%) in an older group. These differences in LOH frequency in these two age groups were significant for chr. 1p21.3 (p = 0.025) only. These data suggest that the deletion pattern in early onset breast tumors is not fully identical to late onset breast tumors. Similar differential deletion patterns of LOH in the 5 highly deleted regions were seen in premenopausal and postmenopausal groups. An association was seen between LOH at chr. 1p34.2 and chr. 1q22-23.2 and higher grade of the tumors in older women. Among the highly deleted regions, the deletion at chr. 1p36 was found to occur early in both groups because of common allelic loss in the bilateral tumors.     

Deletion mapping of the β-glucuronidase gene

GUSB, the gene for β-glucuronidase, has been localized to the proximal long arm of chromosome 7 between 7q11.2 and 7q22. Deficiency of β-glucuronidase results in mucopolysaccharidosis type VII (MPS VII, Sly syndrome). The enzymatic defect has been demonstrated in cultured skin fibroblasts, leukocytes and serum of affected patients. An 8-yr-old boy presented with manifestations similar to MPS VII (mental retardation, short stature, “coarse” facial appearance, mild skeletal involvement and recurrent lower respiratory tract infection) but other, discrepant abnormalities, e. g., bilateral iris colobomata and cleft palate. Normal activity of β-glucuronidase was found in the patient's leukocytes. Chromosome analysis disclosed an interstitial deletion of 7q with one breakpoint at the interface between bands 11.22 and 11.23 and the other breakpoint within band 21.1. DNA from this patient's leukocytes was analyzed for dosage of GUSB sequences. This locus appeared to be present at the normal diploid level. These findings suggest that GUSB is not in the portion of chromosome 7 deleted in our case, narrowing the smallest region of overlap to 7q21.1 → 7q22. We therefore assign the β-glucuronidase gene to 7q21.1 → 7q22.