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Biomedical subjects

A Valent

Publications and source records attributed to A Valent.

At least 19 recordsLinked to original sources
PubMed2005

Conventional and array-based comparative genomic hybridization analysis of nasopharyngeal carcinomas from the Mediterranean area.

Nasopharyngeal carcinoma (NPC) occurs with a high incidence in Southeast Asia and to a lesser extent in the Mediterranean area, especially in Tunisia, Algeria, and Morocco. Cellular gene alterations combined with latent Epstein-Barr virus infection are thought to be essential for NPC oncogenesis. To date, chromosome analysis with comparative genomic hybridization (CGH) has been reported exclusively for NPCs from Southeast Asia. Although NPCs from the Mediterranean area have several distinct clinical and epidemiological features, CGH investigations have been lacking. Chromosome analysis was therefore undertaken on a series of NPC xenografts and biopsies derived from patients of Mediterranean origin. Four xenografts were investigated with a combination of conventional CGH, array-based CGH, and comparative expressed sequence hybridization. In addition, 23 fresh NPC biopsies were analyzed with conventional CGH. Data obtained from xenografts and fresh biopsies were consistent, except that amplification of genes at 18p was observed only in xenografts derived from metastatic tissues. Frequent gains associated with gene overexpression were detected at 1q25 approximately qter (64%) and 12p13 (50%). Losses were noticed mainly at 11q14 approximately q23 (50%), 13q12 approximately q31 (50%), 14q24 approximately q31 (43%), and 3p13 approximately p23 (43%). Comparison with previous reports suggests that Mediterranean NPCs have higher frequencies of gains at 1q and losses at 13q than their Asian counterparts.

Adolescent↗
PubMed2004

No topoisomerase I alteration in a neuroblastoma model with in vivo acquired resistance to irinotecan.

CPT-11 (irinotecan) is a DNA-topoisomerase I inhibitor with preclinical activity against neuroblastoma (NB) xenografts. The aim was to establish in vivo an NB xenograft resistant to CPT-11 in order to study the resistance mechanisms acquired in a therapeutic setting. IGR-NB8 is an immature NB xenograft with MYCN amplification and 1p deletion, which is sensitive to CPT-11. Athymic mice bearing advanced-stage subcutaneous tumours were treated with CPT-11 (27 mg kg(-1) day(-1) x 5) every 21 days (1 cycle) for a maximum of four cycles. After tumour regrowth, a new in vivo passage was performed and the CPT-11 treatment was repeated. After the third passage, a resistant xenograft was obtained (IGRNB8-R). The tumour growth delay (TGD) was reduced from 115 at passage 1 to 40 at passage 4 and no complete or partial regression was observed. After further exposure to the drug, up to 28 passages, the resistant xenograft was definitively established with a TGD from 17 at passage 28. Resistant tumours reverted to sensitive tumours after 15 passages without treatment. IGR-NB8-R remained sensitive to cyclophosphamide and cisplatin and cross-resistance was observed with the topoisomerase I inhibitor topotecan. No quantitative or qualitative topoisomerase I modifications were observed. The level of expression of multidrug resistance 1 (MDR1), MDR-associated protein 1 (MRP1) and, breast cancer resistance protein, three members of the ATP-binding cassette transporter family was not modified over passages. Our results suggest a novel resistance mechanism, probably not involving the mechanisms usually observed in vitro.

Abdominal Neoplasms↗
PubMed2004

Complex translocation (8;12;21): a new variant of t(8;21) in acute myeloid leukemia.

Variants of the t(8;21)(q22;q22) involving chromosomes 8, 21, and other chromosomes account for approximately 3% of all t(8;21)(q22;q22) in acute myeloid leukemia (AML) patients. In this paper, we report a case of AML-M2 with a t(8;12;21)(q22;p12 approximately p13;q22) associated with chromosomal abnormalities, including loss of the Y chromosome and trisomy 8q22 approximately qter. Using a dual-color fluorescence in situ hybridization (FISH) analysis with ETO and AML1 probes, we demonstrated an ETO/AML1 fusion signal on the derivative chromosome 8. Using whole painting probes for chromosomes 8 and 12, we demonstrated a three-way translocation, t(8;12;21)(q22;p12 approximately p13;q22). Reverse transcription polymerase chain reaction (RT-PCR) analysis showed the presence of AML1/ETO fusion transcript. The present case highlights the importance of the combination of approaches, i.e., standard karyotyping, FISH, and RT-PCR, for the detection of variants of t(8;21)(q22;q22), shedding light on region 8q22 approximately qter which could harbor potential genes responsible for leukemogenesis.

Adult↗
PubMed2003

Quality assessment of genetic markers used for therapy stratification.

PURPOSE: Therapy stratification based on genetic markers is becoming increasingly important, which makes commitment to the highest possible reliability of the involved markers mandatory. In neuroblastic tumors, amplification of the MYCN gene is an unequivocal marker that indicates aggressive tumor behavior and is consequently used for therapy stratification. To guarantee reliable and standardized quality of genetic features, a quality-assessment study was initiated by the European Neuroblastoma Quality Assessment (ENQUA; connected to International Society of Pediatric Oncology) Group. MATERIALS AND METHODS: One hundred thirty-seven coded specimens from 17 tumors were analyzed in 11 European national/regional reference laboratories using molecular techniques, in situ hybridization, and flow and image cytometry. Tumor samples with divergent results were re-evaluated. RESULTS: Three hundred fifty-two investigations were performed, which resulted in 23 divergent findings, 17 of which were judged as errors after re-evaluation. MYCN analyses determined by Southern blot and in situ hybridization led to 3.7% and 4% of errors, respectively. Tumor cell content was not indicated in 32% of the samples, and 11% of seemingly correct MYCN results were based on the investigation of normal cells (eg, Schwann cells). Thirty-eight investigations were considered nonassessable. CONCLUSION: This study demonstrated the importance of revealing the difficulties and limitations for each technique and problems in interpreting results, which are crucial for therapeutic decisions. Moreover, it led to the formulation of guidelines that are applicable to all kinds of tumors and that contain the standardization of techniques, including the exact determination of the tumor cell content. Finally, the group has developed a common terminology for molecular-genetic results.

Biomarkers, Tumor↗
PubMed2003

Blastoid mantle cell lymphoma: evidence for nonrandom cytogenetic abnormalities additional to t(11;14) and generation of a mouse model.

Mantle cell lymphoma (MCL) is characterized by the t(11;14)(q13;q32), which is associated with cyclin D1 hyperexpression and a poor prognosis. MCL cases have been shown to progress to a more aggressive disease but the molecular events responsible of this phenomenon have not been determined. We have established two cell lines from the pleural effusions of two patients with MCL that we have used for further cytogenetic characterization to better define the incidence and nature of secondary chromosome abnormalities using multicolor fluorescence in situ hybridization, whole chromosome paint, and specific probes. Both cell lines grew independently without growth factors. Using CCND1/IGH-specific probes, patient UPN1 was found to have a masked t(11;14). Numerous and complex chromosomal abnormalities were found in both cell lines affecting chromosomes 2, 8, 13, 18, 22, X, and Y. These abnormalities included 8p losses, suggesting the presence of an anti-oncogene in this region, rearrangements of 8q24, MYC gene, and translocations involving 8, X, and Y chromosomes, which might be significant in the pathogenesis of MCL progression. The use of the cell lines (UPN1) allowed us to generate a mouse model of human MCL, mimicking a disseminated lymphoma and leading to the death of the animals in 4 weeks. This blastoid MCL model could be of major interest to determine molecular events involved in MCL progression, allowing isolation of involved genes and their functional characterization, and to study the effects of new chemotherapy regimens in mouse models.

Animals↗
PubMed2002

Lack of interstitial chromosome 1p deletions in clinically-detected neuroblastoma.

Loss of heterozygosity (LOH) of the distal part of the short arm of chromosome 1 in neuroblastoma is a well characterised phenomenon. In addition, previous reports have described interstitial deletions outside the common region of loss on chromosome 1p36, suggesting additional tumour suppressor loci. In this study, we have searched extensively for interstitial 1p deletions in a panel of 67 neuroblastoma samples from clinically-detected cases. We used three VNTR probes and 10 dinucleotide markers from the 1p32-36 regions reported to show interstitial deletions. Fifteen (22%) tumours showed telomeric LOH without evidence for more proximal interstitial deletions. Forty-five tumours showed no LOH or allelic imbalance. Seven (10%) tumours demonstrated allelic imbalance for one or more markers. These tumours were subsequently analysed by fluorescent in situ hybridisation (FISH) and flow cytometry. The patterns found in all seven tumours were consistent with copy number changes of the entire chromosome 1, without evidence for interstitial deletions. This study indicates that interstitial deletions of chromosome 1p are rare in clinically-detected neuroblastoma when analysed by a combination of molecular and cytogenetic techniques.

Adolescent↗
PubMed2001

Stromal cells and human malignant neuroblasts derived from bone marrow metastasis may share common karyotypic abnormalities: the case of the IGR-N-91 cell line.

BACKGROUND: Stage IV neuroblastoma is characterized by tumor invasion and metastatic dissemination. Cell lines derived from such neuroblastomas have a high in vitro proliferation capacity. PROCEDURE: We established three neuroblastoma cell lines derived from involved bone marrow of three patients with stage IV neuroblastoma and performed a cytogenetic study. RESULTS: Various culture conditions allowed us to distinguish two cell subpopulations: malignant neuroblasts (Nb-type) and substrate-adherent stromal cells (Str-type). Karyotypic analyses revealed two specific chromosomal abnormalities in diploid malignant IGR-N-331 neuroblasts, der(1)t(1;7)(p22;q11) and der(5)t(5;17)(q35;q21), one unbalanced translocation der(1)t(1;17)(p35;q21)x2 in hyperdiploid malignant IGR-N-337 neuroblasts, and a normal karyotype in both corresponding stromal subpopulations. In contrast, in the IGR-N-91 model, both cell types shared two unbalanced translocations, t(1;4)(q12;p15) and t(2;10)(p14;q11), suggesting that stromal cells and malignant neuroblasts originate from a common stem cell. CONCLUSIONS: Based on our findings, we postulate that genetically modified stromal cells may influence the metastatic potential of malignant neuroblasts.

Bone Marrow Cells↗
PubMed2001

In vivo elimination of acentric double minutes containing amplified MYCN from neuroblastoma tumor cells through the formation of micronuclei.

Neuroblastoma, the most common solid extracranial neoplasm in children, shows an appreciable variability in clinical evolution. Amplification of the MYCN oncogene in this tumor is detected in 25 to 30% of cases and is associated with poor clinical outcome. In this study, quantitative polymerase chain reaction and fluorescence in situ hybridization were used to determine MYCN amplification status in 46 neuroblastoma tumors. MYCN amplification was detected in tumors from 11 patients. Fluorescence in situ hybridization revealed the presence of micronuclei containing amplified MYCN sequences in 8 of the 11 tumors. Micronuclei are indicative of spontaneous elimination or loss of amplified sequences by tumor cells. Because the elimination of amplified sequences can be enhanced in vitro by specific drugs such as hydroxyurea, our observations suggest a new therapeutic strategy specifically targeted to cells with amplified genes.

Child, Preschool↗
PubMed2000

[Detection of secretion of beta-galactosidase by plant cells].

A simple, sensitive and reproducible method of detection of extracellular beta-galactosidase was worked out. beta-Galactosidase secreted by the callus culture, or the roots of sprouting plants, hydrolyzes the substrate (6-bromo-2-naphthyl-beta-D-galactopyranoside) to produce beta-D-galactose and 6-bromo-2-naphthol, which after simultaneous azocoupling with hexazotized p-rosaniline, or basic fuchsine, produce a reddish brown compound, nearly insoluble in water (a diazonium salt). The colouring under the callus culture and around it, as well as the colouring of the roots of the sprouting plants, is a manifestation of the activity of extracellular beta-galactosidase by the objects under study. The intensity of the colouring is a measure of their enzymatic activity. The share of intracellularly localized enzyme represents the majority and the share of extracellular beta-galactosidase the minority.

Plant Roots↗
PubMed1999

Purification of a novel flavoprotein involved in the thyroid NADPH oxidase. Cloning of the porcine and human cdnas.

Hydrogen peroxide is the final electron acceptor for the biosynthesis of thyroid hormone catalyzed by thyroperoxidase at the apical surface of thyrocytes. Pig and human thyroid plasma membrane contain a Ca(2+)-dependent NAD(P)H oxidase that generates H(2)O(2) by transferring electrons from NAD(P)H to molecular oxygen. We purified from pig thyroid plasma membrane a flavoprotein which constitutes the main, if not the sole, component of the thyroid NAD(P)H oxidase. Microsequences permitted the cloning of porcine and human full-length cDNAs encoding, respectively, 1207- and 1210-amino acid proteins with a predicted molecular mass of 138 kDa (p138(Tox)). Human and porcine p138(Tox) have 86.7% identity. The strongest similarity was to a predicted polypeptide encoded by a Caenorhabditis cDNA and with rbohA, a protein involved in the Arabidopsis NADPH oxidase. p138(Tox) shows also similarity to the p65(Mox) and to the gp91(Phox) in their C-terminal region and have consensus sequences for FAD- and NADPH-binding sites. Compared with gp91(Phox), p138(Tox) shows an extended N-terminal containing two EF-hand motifs that may account for its calcium-dependent activity, whereas three of four sequences implicated in the interaction of gp91(Phox) with the p47(Phox) cytosolic factor are absent in p138(Tox). The expression of porcine p138(Tox) mRNA analyzed by Northern blot is specific of thyroid tissue and induced by cyclic AMP showing that p138(Tox) is a differentiation marker of thyrocytes. The gene of human p138(Tox) has been localized on chromosome 15q15.

Amino Acid Sequence↗
PubMed1999

Phenotypic and genotypic diversity of human neuroblastoma studied in three IGR cell line models derived from bone marrow metastases.

Metastatic stage IV neuroblastoma tumors, as well as cell lines derived from them, are highly malignant and rapidly fatal. To determine whether malignant potential of these cells might be influenced by stromal tissue at sites frequently involved in metastasis, we initiated primary cultures from bone marrow of three patients (331, 337, and 91) with stage IV neuroblastoma. All three explants contained two distinct cell populations, malignant neuroblasts (Nb-type) and substrate adherent stromal-like (Str-type) cells. The cell types were separated at the first passage and studied by cytogenetic, molecular, and immunocytochemical methods. Karyotypic analyses after 3-6 passages in vitro revealed the presence of unique chromosomal abnormalities in Nb-type cells of all three lines: (1) der(1)t(1;7) (p32;q11) and der(5)t(5;17)(q35;q21) in pseudodiploid IGR-N-331 neuroblasts; (2) der(1)t(1;17)(p35;q21-22) x 2 and der(7)t(7;7)(p21;q21) in IGR-N-337 hyperdiploid neuroblasts; and (3) more than six rearranged chromosomes in two related subpopulations of hypodiploid IGR-N-91 neuroblasts. Neuroblastic cells from all three tumors amplified MYCN 25- to 50-fold (with amplified genes visible as dmin or, in one IGR-N-91 subline, as an hsr(14)[q32]) and expressed N-CAM. Str-type cells from tumors 331 and 337 had a normal diploid karyotype, did not express either N-CAM or S-100, and are probably normal bone marrow fibroblasts. By contrast, S-100 negative Str-type IGR-N-91 cells were hypodiploid and shared at least two unbalanced translocations, der(4)t(1;4)(q12;p15) and der(2)t(2;10;17)(p14;q11;q22), with neuroblastic counterparts, indicating that "stromal" cells and malignant neuroblasts had a common tumor cell origin. Thus, the Str-type cells of IGR-N-91 are examples of S-type phenotypic variants frequently described for long-term human neuroblastoma cells lines in vitro, but not previously observed in vivo.

Bone Neoplasms↗
PubMed1999

Thiohydrazone copper(II) complexes. The relationship between redox properties and superoxide dismutase mimetic activity.

The redox behaviour of copper(II) complexes with the open chain ligand, benzilbisthiosemicarbazone, and the macrocyclic one [3,4,10,11-tetraphenyl-1,2,5,8,9,12,13-octaazacyclotetradeca-7,14- dithione- 2,4,9,11-tetraene] has been explored by cyclic voltammetry. The half-wave potential values for the copper(II)/copper(I) redox couple and the spectral data obtained on dimethylsulfoxide (DMSO) solution agree with the superoxide dismutase (SOD)-mimetic activity of the complexes. The macrocyclic complexes show more positive reduction potential and more activity than the open chain derivatives. From our results it follows that the structure and conformation of ligand has influence on the redox potential of central atom in coordination compound. The changes in the coordination sphere are connected with the change of biological function of compounds represented by SOD-mimic activity. In addition, the L1H6 derivatives show quasireversible waves associated to Cu(II)/Cu(III) process.

Binding Sites↗
PubMed1997

Monoallelically expressed gene related to p53 at 1p36, a region frequently deleted in neuroblastoma and other human cancers.

We describe a gene encoding p73, a protein that shares considerable homology with the tumor suppressor p53. p73 maps to 1p36, a region frequently deleted in neuroblastoma and other tumors and thought to contain multiple tumor suppressor genes. Our analysis of neuroblastoma cell lines with 1p and p73 loss of heterozygosity failed to detect coding sequence mutations in remaining p73 alleles. However, the demonstration that p73 is monoallelically expressed supports the notion that it is a candidate gene in neuroblastoma. p73 also has the potential to activate p53 target genes and to interact with p53. We propose that the disregulation of p73 contributes to tumorigenesis and that p53-related proteins operate in a network of developmental and cell cycle controls.

Alleles↗
PubMed1996

Assignment of the NTRK4 (trkE) gene to chromosome 6p21.

Reverse transcriptase-polymerase chain reactions using foetal brain RNA with reverse and forward primers of the first, second and third NTRK4 region allowed us to obtain three amplified NTRK4 fragments. The specificity of amplified fragments was checked by digestion with restriction endonucleases AvrII, HindIII and PspII for the first, second and third regions, respectively. Each restriction site was specific for each amplified fragment. The fragment of the NTRK4 first region was also sequenced and the sequence determined was identical to the human NTRK4 sequence. The three amplified fragments were cloned in pBS. For the Southern technique, plasmid pBS-NTRK4a (with an insert of 1052 bp) detected a human 9-kb HindIII sequence which was localised unambiguously on chromosome 6. For fluorescence in situ hybridisation, the three plasmids, pBS-NTRK4a, pBS-NTRK4b (insert 924 bp) and pBS-NTRK4c (insert 1114 bp) were pooled and used as a probe. This NTRK4 probe was localised on 6p21. Of 50 metaphases analysed, 49 contained twin spot signals on both sister chromatids.

Base Sequence↗
PubMed1996

MDM2 amplification in a primary alveolar rhabdomyosarcoma displaying a t(2;13)(q35;q14).

This report describes a case of rhabdomyosarcoma associated with a 2;13 translocation and multiple double minute chromosomes. The origin of the amplified DNA was identified using comparative genomic hybridization, which pinpointed a unique spot at 12q13-->q14. Band 12q13 has been shown to contain several genes that are occasionally amplified in other sarcomas. Fluorescene in situ hybridization to tumor metaphases with probes specific for this region indicated that the double minutes contained the MDM2 gene but not the CDK4 gene. MDM2 amplification was further quantified by Southern hybridization, which showed a mean value of 25 copies per haploid genome. This is the first example of MDM2 amplification in a rhabdomyosarcoma.

Adolescent↗
PubMed1996

Variant forms of Philadelphia translocation in two patients with chronic myeloid leukemia.

During a 4-year period (December 1990-December 1994), among other diagnoses one hundred cases of chronic myeloid leukemia (CML) were analyzed in our department. We focused our attention on two cases with a variant form of Philadelphia translocation. Cytogenetic and molecular genetic studies were performed to resolve the status of BCR and ABL in the bone marrow or peripheral blood cells of the two CML patients with complex translocations involving chromosomes 3, 9, 22 and 9, 12, 22 respectively. In the first case the presence of Ph chromosome was detected cytogenetically, BCR-ABL translocation was detected by Southern hybridization. In the second case, only the PCR method showed BCR-ABL rearrangement. The second case, with a random variant form of Ph translocation, could be detected using different methods of clinical molecular genetics.

Adult↗
PubMed1996

[Rapid identification of chromosomes by in situ hybridization of labelled oligonucleotides and comparison with the PRINS method].

We propose a simple, fast and inexpensive method of identification of human centromeres on metaphasic chromosomes and interphasic nuclei. This is based on in situ hybridization of labelled oligonucleotides. The efficiency of the methodology was demonstrated on cytogenetic preparations from human heteroploid and human x hamster hybrid cell lines and also on frozen tissue sections using an oligonucleotide specific for the alpha-satellite DNA of chromosome 1. Three versions of this oligonucleotide respectively labelled with 1, 4 and 10 fluorescein molecules were synthesized. The signal intensity provided by the oligonucleotide coupled with 4 fluoresceins allowed unambiguously the detection of the chromosome and the establishment of its ploidy using a classical cytogenetic microscope without the need for an amplification procedure. The use of different fluorochromes and possibly combination with an unlabelled elongation in 3' of the oligonucleotides which stabilize its hybridization, lead to a simple multicolour method. Preliminary quantification of the signals obtained by in situ hybridization of labelled oligonucleotides and comparison with those obtained by primed in situ labelling (PRINS) using the same nucleotides as primers, suggest that the elongation generated by PRINS may be very short compared with a PCR in solution. This limited efficiency of the in situ elongation may reflect the present difficulties of PRINS and DISC PCR (direct in situ single copy polymerase chain reaction) with primers specific for non-repetitive sequencies.

Animals↗