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R L Stallings

Publications and source records attributed to R L Stallings.

At least 19 recordsLinked to original sources
PubMed2004

Molecular cytogenetic analysis of recurrent unbalanced t(11;17) in neuroblastoma.

Loss of 11q material occurs in approximately 30% of advanced stage neuroblastoma and defines a distinct genetic subtype of this disease. These tumors almost always possess unbalanced gain of the 17q, along with many additional recurrent chromosomal imbalances. Loss of 11q and gain of 17q is often the consequence of an unbalanced translocation between the long arms of both chromosomes, but because of the involvement of other chromosomal mechanisms, the actual frequency of t(11;17) is unknown. In addition, chromosomal breakpoint positions for the t(11;17) are variable in different tumors, with breakpoints on neither the 11q nor 17q being well defined. We have used interphase fluorescence in situ hybridization analysis to detect a der(11)t(11;17) in a series of neuroblastomas with 11q loss/17q gain using a statistical approach which could be applicable to the detection of translocations in other solid tumors. The frequency of der(11)t(11;17) was approximately 90% in our neuroblastoma series. A balanced t(11;17) was also detected in a MYCN amplified tumor, which is a distinctly different genetic subtype from the 11q- tumors. Breakpoint positions on 11q were determined to be variable, whereas all breakpoints on 17q appeared to cluster proximal to position 43.1 Mb on the DNA sequence map. The majority of tumors had large numbers of nuclei with 2 or more copies of der(11)t(11;17), which led to unbalanced gain of 11p, and further increases in 17q imbalance. The prevalence of t(11;17) in neuroblastoma warrants additional studies to further define the range in variation in breakpoint positions on both chromosomes and to elucidate the molecular mechanisms that lead to this important and interesting recurrent genetic abnormality.

Chromosomes, Artificial, Bacterial↗
PubMed2004

Oligonucleotide microarray analysis of gene expression in neuroblastoma displaying loss of chromosome 11q.

A number of distinct subtypes of neuroblastoma exist with different genetic abnormalities that are predicative of outcome. Whole chromosome gains are usually associated with low stage disease and favourable outcome, whereas loss of 1p, 3p and 11q, unbalanced gain of 17q and MYCN amplification (MNA) are indicative of high stage disease and unfavourable prognosis. Although MNA and loss of 11q appear to represent two distinct genetic subtypes of advanced stage neuroblastoma, a detailed understanding of how these subtypes differ in terms of global gene expression is still lacking. We have used metaphase comparative genomic hybridization (CGH) analysis in combination with oligonucleotide technology to identify patterns of gene expression that correlate with specific genomic imbalances found in primary neuroblastic tumours and cell lines. The tumours analysed in this manner included a ganglioneuroma, along with various ganglioneuroblastoma and neuroblastoma of different stages and histopathological classifications. Oligonucleotide microarray-based gene expression profile analysis was performed with Affymetrix HU133A arrays representing approximately 14 500 unique genes. The oligonucleotide microarray results were subsequently validated by quantitative real-time PCR, immunohistochemical staining, and by comparison of specific gene expression patterns with published results. Hierarchical clustering of gene expression data distinguished tumours on the basis of stage, differentiation and genetic abnormalities. A number of genes were identified whose patterns of expression were highly correlated with 11q loss; supporting the concept that loss of 11q represents a distinct genetic subtype of neuroblastoma. The implications of these results in the process of neuroblastoma development and progression are discussed.

Biomarkers, Tumor↗
PubMed2004

Evolution of unbalanced gain of distal chromosome 2p in neuroblastoma.

Neuroblastoma, one of the most common tumors of childhood, presents at diagnosis with a vast number of recurrent chromosomal imbalances that include hyperdiploidy for whole chromosomes, partial loss of 1p, 3p, 4p, 11q, 14q, partial gain of 1q, 7q, 17q and amplification of MYCN. These abnormalities are nonrandomly distributed in neuroblastoma as loss of 3p and 11q rarely occur in MYCN amplified neuroblastomas. Here, we report on a patient who had a non-MYCN amplified 3p-/11q- neuroblastoma at diagnosis who subsequently developed a high level of MYCN amplification in bone marrow metastases 41 months after induction of complete remission. The tumor at diagnosis had low level unbalanced gain of distal 2p. In order to assess the frequency of low level gain of distal 2p in neuroblastoma, we examined the comparative genomic hybridization results from 60 neuroblastomas. Among non-MYCN amplified neuroblastomas, 8/45 (18%) had low level gain of distal 2p. Low level gain for a segment of 2p (i.e. a region larger than the 2p23-->p24 undergoing amplification) was also detected in five of the 15 tumors that had high level MYCN amplification. The possibility that low level gain of distal 2p is a risk factor for high level MYCN amplification is discussed.

Abdominal Neoplasms↗
PubMed2003

Are gains of chromosomal regions 7q and 11p important abnormalities in neuroblastoma?

Neuroblastoma exhibiting deletion of a segment of the long arm of chromosome 11 represents a genetic subtype of tumor that is distinct from those exhibiting MYCN amplification or 1p deletion. The 11q- genetic subtype is further characterized by gain of 17q and loss of distal 3p material. Gain of 11p material has also been reported in neuroblastoma with 11q loss, but at a considerably lower frequency than gain of 17q or loss of the distal 3p region. Our results, however, indicate that gain of 11p may occur more frequently in 11q- neuroblastoma than what was previously realized. Comparative genomic hybridization analyses of neuroblastoma tissue from eleven patients indicated that six of 11 tumors (55%) with loss of 11q also possessed gain of 11p. The shortest region of 11p gain was 11p11.2-->p14. G-banding and fluorescence in situ hybridization analysis performed on tumor cells from primary and metastatic sites from one patient allowed us to infer that gain of 11p arose secondarily to the abnormality that led to the loss of 11q material. Gain of an entire chromosome 7 was detected in 17 of 43 (40%) tumors, whereas gain of 7q was detected in 5 of 43 (12%) tumors. Unlike gain of 11p, gain of an entire chromosome 7 appears to be prevalent in all tumor stages and is not limited to the 11q- tumor subtype. Gain of 7q, however, is more prevalent in higher stage tumors. G-band cytogenetic analysis indicated that an unbalanced t(3;7) was responsible for the gain of 7q and loss of 3p material in one case. We discuss the possibility that gain of 7/7q, and 11p material may contribute to either tumorigenesis or progression.

Abdominal Neoplasms↗
PubMed2001

Genetic abnormalities in a pre and post-chemotherapy hepatoblastoma.

Comparative genomic hybridization (CGH) analysis was performed on both a pre- and post-chemotherapy hepatoblastoma from a 24-month-old female patient. The diagnostic sample obtained from a tru-cut biopsy was a mixed epithelial-mesenchymal tumor with both fetal and embryonal patterns present. In contrast, the post-chemotherapy tumor exhibited a prominent anaplastic large cell population focally reminiscent of pleomorphic hepatocellular carcinoma (HCC). CGH analysis indicated that there were similarities as well as differences in the gains and losses of genetic material in each tumor. The diagnostic sample had gains of chromosome 1q, 2, 2(q31q33), 7, 8q, 12(q15q22), 17q and 20 material, while the post-chemotherapy tumor had gains of 1q, 2, 7, 8q, 10, 17q and 20 material. In addition, the pre- and post-chemotherapy samples may have incurred loss of chromosome 17p material. The main differences between the two samples involved localized gain of 2(q31q33) and 12(q15q22) in the pre-chemotherapy sample, and gain of chromosome 10 material in the post-chemotherapy tumor. The patient subsequently developed metastatic nodules in her lungs, the histology of which was identical in pattern to the diagnostic pattern, and appeared to have localized gain of 2(q31q33) and 12(q15q22). These results are consistent with published results that gain of chromosome 8q and 20 are associated with an unfavorable prognosis.

Child, Preschool↗
PubMed2000

Coordinate deletion of chromosome 3p and 11q in neuroblastoma detected by comparative genomic hybridization.

Neuroblastoma, the most common extracranial solid tumor of childhood, is associated with a number of genetic abnormalities that are prognostically significant. The most common abnormalities are associated with aggressive clinical behavior and include deletion of distal chromosome 1p, NMYC amplification, and unbalanced gain of the long arm of chromosome 17. There are also other recurrent, but less frequent abnormalities, the clinical significance of which is uncertain. These less common abnormalities include deletion 3p, 11q, and 14q. To gain further clinical insight into some of the less commonly observed abnormalities in neuroblastoma, we performed comparative genomic hybridization (CGH) analysis on 24 primary and metastatic neuroblastomas (6 stage 2, 5 stage 3, 11 stage 4, and 2 stage 4). Nineteen of these tumors were prechemotherapy. A total of 190 abnormalities were detected from these tumors. Four of the 24 tumors studied showed loss of 11q material, with 3 of these tumors also possessing distal chromosome 3p deletions. Our results provide confirmation that deletion of chromosome 3p is nonrandomly associated with deletion of chromosome 11q in neuroblastoma. However, analysis of our results, along with other results reported in the literature, indicate that there is no statistically significant association between 3p and 11q loss and more clinically aggressive tumors.

Abdominal Neoplasms↗
PubMed2000

Comparative genomic hybridization in pediatric acute lymphoblastic leukemia.

Comparative genomic hybridization (CGH) was used to clarify the chromosomal status of 15 patients diagnosed with childhood acute lymphoblastic leukemia (ALL). Bone marrow samples from 10 of the 15 patients were selected because no metaphases were obtained for cytogenetic analysis. Three patients with normal trypsin and giemsa banding (GTG) karyotypes were also studied by CGH to determine whether significant abnormalities might have been missed by banding analysis, and samples from an additional 2 patients with hyperdiploidy were also included. Seven of the 10 patients with failed GTG banding analysis were found to be chromosomally abnormal by CGH; 2 out of 3 patients with normal GTG band karyotypes were abnormal, indicating that the metaphases available for karyotyping were not malignant cells, and that CGH analysis of hyperdiploid samples provided more accurate resolution than karyotyping alone. The prognostic value of chromosomal aberrations detected by CGII and the efficiency of the technique suggest a central role for CGH in routine clinical cytogenetics.

Bone Marrow↗
PubMed1999

Applications of comparative genomic hybridisation in constitutional chromosome studies.

G band cytogenetic analysis often leads to the discovery of unbalanced karyotypes that require further characterisation by molecular cytogenetic studies. In particular, G band analysis usually does not show the chromosomal origin of small marker chromosomes or of a small amount of extra material detected on otherwise normal chromosomes. Comparative genomic hybridisation (CGH) is one of several molecular approaches that can be applied to ascertain the origin of extra chromosomal material. CGH is also capable of detecting loss of material and thus is also applicable to confirming or further characterising subtle deletions. We have used comparative genomic hybridisation to analyse 19 constitutional chromosome abnormalities detected by G band analysis, including seven deletions, five supernumerary marker chromosomes, two interstitial duplications, and five chromosomes presenting with abnormal terminal banding patterns. CGH was successful in elucidating the origin of extra chromosomal material in 10 out of 11 non-mosaic cases, and permitted further characterisation of all of the deletions that could be detected by GTG banding. CGH appears to be a useful adjunct tool for either confirming deletions or defining their breakpoints and for determining the origin of extra chromosomal material, even in cases where abnormalities are judged to be subtle. We discuss internal quality control measures, such as the mismatching of test and reference DNA in order to assess the quality of the competitive hybridisation effect on the X chromosome.

Chromosome Banding↗
PubMed1995

Primary structure of human lumican (keratan sulfate proteoglycan) and localization of the gene (LUM) to chromosome 12q21.3-q22.

A human corneal fibroblast cDNA library was screened with a bovine lumican cDNA probe to obtain three clones. Sequencing of the longest clone (1.75 kb) yielded an open reading frame of 1014 bp coding for a 338-amino-acid core protein. Amino acid sequencing of a tryptic peptide resulted in a 9-amino-acid match with the derived primary structure, confirming the identity of these clones. Human lumican displays all of the features of small interstitial proteoglycans: N- and C-terminal domains with highly conserved cysteines and a central domain containing nine repeats of slight variations of the leucine motif LXXLXLXXNXL. Like bovine lumican, the human core protein contains four possible N-glycosylation sites in the central domains, all or some of which are substituted with keratan sulfate side chains. At the amino acid level, it is 90% identical with bovine and 72% identical with the chicken core protein. The gene (LUM) was localized to human chromosome 12 by hybridizing a cDNA probe to a Southern blot containing a human/hamster monochromosomal mapping panel DNA. Further sublocalization to 12q21.3-q22 was performed by the fluorescence in situ hybridization technique using a lumican P1 genomic clone. By immunohistochemical staining, we show lumican's presence, not only in the corneal stroma as shown previously, but also in the dermal area of the skin, indicating a wider distribution of this proteoglycan.

Amino Acid Sequence↗
PubMed1995

Efficient pooling designs for library screening.

We describe efficient methods for screening clone libraries, based on pooling schemes that we call "random k-sets designs." In these designs, the pools in which any clone occurs are equally likely to be any possible selection of k from the v pools. The values of k and v can be chosen to optimize desirable properties. Random k-sets designs have substantial advantages over alternative pooling schemes: they are efficient, flexible, and easy to specify, require fewer pools, and have error-correcting and error-detecting capabilities. In addition, screening can often be achieved in only one pass, thus facilitating automation. For design comparison, we assume a binomial distribution for the number of "positive" clones, with parameters n, the number of clones, and c, the coverage. We propose the expected number of resolved positive clones--clones that are definitely positive based upon the pool assays--as a criterion for the efficiency of a pooling design. We determine the value of k that is optimal, with respect to this criterion, as a function of v, n, and c. We also describe superior k-sets designs called k-sets packing designs. As an illustration, we discuss a robotically implemented design for a 2.5-fold-coverage, human chromosome 16 YAC library of n = 1298 clones. We also estimate the probability that each clone is positive, given the pool-assay data and a model for experimental errors.

Binomial Distribution↗
PubMed1995

Conservation and evolution of (CT)n/(GA)n microsatellite sequences at orthologous positions in diverse mammalian genomes.

The distribution and evolution of (CT)n microsatellites were examined in GenBank mammalian DNA sequences because these microsatellites are known to play important roles in the regulation of some genes in Drosophila melanogaster. A total of 236 (CT)n microsatellite loci were found in GenBank mammalian gene sequences. To determine whether (CT)n microsatellite arrays were conserved at orthologous positions in distantly related mammalian sequences, we determined whether orthologous sequences existed in GenBank for each of the 236 loci. A total of 47 sequence alignments could be made. For rodent x rodent comparisons, 7 of 8 (CT)n arrays were conserved at identical positions in each pair of orthologous sequences. Comparisons of orthologous sequences between different orders of mammals indicated that 11 of 39 (CT)n arrays occurred at orthologous positions or within 1 kb of orthologous positions in each pair of sequences. It appears that there is some level of conservation of (CT)n repeats in distantly related mammals. However, this level of conservation may not be greater than what might be expected to occur by chance. In 13 cases where (CT)n arrays were not conserved at orthologous positions, the lack of a (CT)n array in one sequence resulted from either nucleotide substitution within an array or nonexpansion of a shorter (CT)n element. In these cases, significant sequence identity could be detected throughout the entire region even though the repeat array was not detected in one of the sequences. In contrast, there was a disruption of sequence identity in the (CT)n microsatellite region that ranged from 24 to 1600 bp in 21 cases.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

A large duplicated area in the polycystic kidney disease 1 (PKD1) region of chromosome 16 is prone to rearrangement.

An area of 500 kb at the proximal end of the polycystic kidney disease 1 (PKD1) region has been mapped in detail, with 260 kb cloned in cosmids. The area cloned from normal individuals contains two homologous but divergent regions each of 75 kb, including the previously described marker 26-6. Pulsed-field gel electrophoresis identified a duplication of 75 kb of this region, referred to as the OX duplication (OXdup), in three patients with PKD1. The OXdup probably arose by an unequal exchange promoted by misalignment of partially homologous areas. Study of the OXdup in a large PKD1 family showed that it segregated with PKD1 in just one-half of the family, indicating that a recent crossover had occurred between the OXdup and PKD1 and showing that it was not a PKD1 mutation. Further analysis identified an OXdup breakpoint fragment: the OXdup was subsequently identified in 2 normal individuals of 110 assayed. The finding of the OXdup and in other individuals an 11-kb deletion (OXdel) at a similar point within this duplicated area indicates that this is an unusually unstable genomic region.

Chromosome Mapping↗
PubMed1994

FISH mapping of a human chromosome 16 constitutional pericentric inversion inv(16)(p13q22) found in a large kindred.

Fluorescence in situ hybridization analysis (FISH) was used to map the constitutional chromosome 16 pericentric inversion breakpoints inv(16)(p13q22) detected in one individual (II-2) from a large kindred [Bianchi et al., 1992: Am J Med Genet 43:791-795]. The breakpoints found in individual II-2 mapped to distinctly different locations than the chromosome 16 pericentric inversion breakpoints commonly acquired in acute nonlymphocytic leukemia. The constitutional pericentric inversion breakpoints also do not map to regions where low abundance repetitive DNA sequences found in bands 16p13 and q22 are located. The results indicate that low abundance, chromosome 16-specific repetitive DNA sequences in bands p13 and q22 are probably not causally related to the inversion that is found in many members of a large kindred [Bianchi et al., 1992].

Adult↗
PubMed1994

Distribution of trinucleotide microsatellites in different categories of mammalian genomic sequence: implications for human genetic diseases.

The distribution of all trinucleotide microsatellite sequences in the GenBank database was surveyed to provide insight into human genetic disease syndromes that result from expansion of microsatellites. The microsatellite motif (CAG)n is one of the most abundant microsatellite motifs in human GenBank DNA sequences and is the most abundant microsatellites found in exons. This fact may explain why (CAG)n repeats are thus far the predominant microsatellites expanded in human genetic diseases. Surprisingly, (CAG)n microsatellites are excluded from intronic regions in a strand-specific fashion, possibly because of similarity to the 3' consensus splice site, CAGG. A comparison of the positions of microsatellites in human vs rodent homologous sequences indicates that some arrays are not extensively conserved for long periods of time, even when they form parts of protein coding sequences. The general lack of conservation of trinucleotide repeat loci in diverse mammals indicates that animal models for some human microsatellite expansion syndromes may be difficult to find.

Animals↗