RESUMEN
Chromosomal translocations represent an important prognostic indicator in B-cell chronic lymphocytic leukemia (B-CLL). However, their value had been neither determined in homogeneously treated patients nor compared to that of IgV(H) mutational status. Sixty-five B-CLL patients were investigated using cytogenetics, interphase fluorescence in situ hybridization (FISH), analysis of IgV(H) and of TP53 mutational status before treatment with 2-chloro-2'-deoxyadenosine (CdA). Translocations (n=45) were detected in 42% of the patients, including both balanced (n=12) and unbalanced (n=33) types. IgV(H) was mutated in 43% of the patients. Patients with translocations were more heavily pretreated (P=0.05), presented with more complex karyotypes (P<0.001), 17p abnormalities and TP53 mutations, and had a higher failure rate (59 vs 21% in patients without translocations, P=0.004). Patients with unbalanced translocations displayed a shorter median treatment-free survival (TFS, 6.9 vs 35.9 months, log rank 22.72, P<0.001) and overall survival (OS, 13.0 vs 68.0 months, log rank 16.51, P<0.001), as compared to patients without translocation. In multivariate analysis, unbalanced translocations were independently associated with therapeutic failure, short TFS and short OS. IgV(H) mutational status was independently associated with risk of failure and TFS, but not OS. In B-CLL patients treated with CdA, translocations are strong predictors of outcome.
Asunto(s)
Antineoplásicos/uso terapéutico , Cladribina/uso terapéutico , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/mortalidad , Translocación Genética , Adulto , Anciano , Anciano de 80 o más Años , Aberraciones Cromosómicas , Cromosomas Humanos Par 11/genética , Cromosomas Humanos Par 13/genética , Cromosomas Humanos Par 17/genética , Femenino , Humanos , Hibridación Fluorescente in Situ , Interfase , Cariotipificación , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Masculino , Persona de Mediana Edad , Mutación/genética , Pronóstico , Factores de Riesgo , Tasa de Supervivencia , Insuficiencia del Tratamiento , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
We report on a 48-year-old man with concomitantly diagnosed kappa expressing chronic lymphocytic leukaemia (CLL) and lambda light chain disease with highly complex chromosomal aberrations. The clinical course of the disease was very aggressive with survival of only 1 month. We demonstrate the distinct clonal origin by cytogenetic data and immunoglobulin rearrangement studies. To our knowledge this is the first report of a light chain disease associated with CLL.
Asunto(s)
Hipergammaglobulinemia/complicaciones , Cadenas Ligeras de Inmunoglobulina , Leucemia Linfocítica Crónica de Células B/complicaciones , Cromosomas Humanos Par 13 , Cromosomas Humanos Par 14 , Resultado Fatal , Reordenamiento Génico de Cadena Pesada de Linfocito B , Reordenamiento Génico de Cadena Ligera de Linfocito B , Humanos , Hipergammaglobulinemia/genética , Cariotipificación , Leucemia Linfocítica Crónica de Células B/genética , Masculino , Persona de Mediana EdadRESUMEN
AIMS: The diagnosis of gastrointesinal stromal tumours (GISTs) is widely based on morphological features and KIT (CD117) immunoreactivity. Most patients with advanced GISTs show a major clinical response after treatment with imatinib mesylate. The histopathological features of GISTs in patients on prolonged imatinib treatment have, thus far, not been addressed in detail. In this report, we present three patients with metastatic GISTs, who received more than 1 year of therapy with imatinib, and whose tumours changed their morphological and immunohistochemical characteristics during continued treatment with the drug. METHODS AND RESULTS: All three primary GISTs from these patients were classical spindle-type tumours, showing diffuse, strong CD117, CD34, and focal alpha-smooth muscle actin expression. During treatment, two clinically progressive and one clinically stable GIST revealed a diffuse epithelioid, or pseudopapillary epithelioid growth pattern, characterized by rounded cells with eosinophilic cytoplasm and uniform round-to-ovoid nuclei. In addition, GIST specimens from patients on therapy showed complete loss of CD117 immunoreactivity. Remarkably, two of these tumours also became CD34 immunonegative and in one case the progression was accompanied by desmin expression. KIT mutational analysis revealed the presence of distinct exon 11 mutant isoforms in all cases examined, while the same genotype was sustained in the base line and on-therapy tumour specimens, proving the common origin of analysed specimens. CONCLUSIONS: GISTs subject to imatinib treatment can undergo striking (immuno)phenotypic changes, which are not necessarily corroborated by new genotypic modifications. Because these may mimic other tumour types, this feature creates a differential diagnostic challenge, of which the pathologist should be aware.
Asunto(s)
Tumores del Estroma Gastrointestinal/tratamiento farmacológico , Tracto Gastrointestinal/efectos de los fármacos , Inmunohistoquímica/normas , Piperazinas/uso terapéutico , Pirimidinas/uso terapéutico , Actinas/análisis , Adulto , Anciano , Antígenos CD34/análisis , Antineoplásicos/uso terapéutico , Benzamidas , Biomarcadores de Tumor/análisis , Desmina/análisis , Tumores del Estroma Gastrointestinal/diagnóstico , Tumores del Estroma Gastrointestinal/metabolismo , Tracto Gastrointestinal/química , Tracto Gastrointestinal/patología , Humanos , Mesilato de Imatinib , Inmunohistoquímica/métodos , Queratinas/análisis , Persona de Mediana Edad , Músculo Liso/química , Proteínas Proto-Oncogénicas c-kit/análisis , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
Although reciprocal chromosomal translocations are not typical for B-cell chronic lymphocytic leukemia (B-CLL), we identified the novel t(1;6)(p35.3;p25.2) in eight patients with this disorder. Interestingly, all cases showed lack of somatically mutated IgV(H). Clinical, morphological, immunologic, and genetic features of these patients are described. Briefly, the age ranged from 33 to 81 years (median: 62.5 years) and the sex ratio was 6M:2F. Most of the patients (6/8) presented with advanced clinical stage. Therapy was required in seven cases. After a median follow-up of 28 months, five patients are alive and three died from disease evolution. Three cases developed transformation into diffuse large B-cell lymphoma. Translocation t(1;6) was found as the primary karyotypic abnormality in three patients. Additional chromosomal aberrations included changes frequently found in unmutated B-CLL, that is, del(11)(q), trisomy 12 and 17p aberrations. Fluorescence in situ hybridization analysis performed in seven cases allowed us to map the t(1;6) breakpoints to the 1p35.3 and 6p25.2 chromosomal bands, respectively. The latter breakpoint was located in the genomic region coding for MUM1/IRF4, one of the key regulators of lymphocyte development and proliferation, suggesting involvement of this gene in the t(1;6). Molecular characterization of the t(1;6)(p35.3;p25.2), exclusively found in unmutated subtype of B-CLL, is in progress.
Asunto(s)
Cromosomas Humanos Par 1 , Cromosomas Humanos Par 6 , Leucemia Linfocítica Crónica de Células B/genética , Translocación Genética , Humanos , Hibridación Fluorescente in Situ , CariotipificaciónRESUMEN
In T-cell acute lymphoblastic leukemia (T-ALL), transcription factors are known to be deregulated by chromosomal translocations, but mutations in protein tyrosine kinases have only rarely been identified. Here we describe the extrachromosomal (episomal) amplification of ABL1 in 5 of 90 (5.6%) individuals with T-ALL, an aberration that is not detectable by conventional cytogenetics. Molecular analyses delineated the amplicon as a 500-kb region from chromosome band 9q34, containing the oncogenes ABL1 and NUP214 (refs. 5,6). We identified a previously undescribed mechanism for activation of tyrosine kinases in cancer: the formation of episomes resulting in a fusion between NUP214 and ABL1. We detected the NUP214-ABL1 transcript in five individuals with the ABL1 amplification, in 5 of 85 (5.8%) additional individuals with T-ALL and in 3 of 22 T-ALL cell lines. The constitutively phosphorylated tyrosine kinase NUP214-ABL1 is sensitive to the tyrosine kinase inhibitor imatinib. The recurrent cryptic NUP214-ABL1 rearrangement is associated with increased HOX expression and deletion of CDKN2A, consistent with a multistep pathogenesis of T-ALL. NUP214-ABL1 expression defines a new subgroup of individuals with T-ALL who could benefit from treatment with imatinib.
Asunto(s)
Genes abl , Leucemia-Linfoma de Células T del Adulto/genética , Proteínas de Complejo Poro Nuclear/genética , Plásmidos/genética , Secuencia de Aminoácidos , Fusión Artificial Génica , Secuencia de Bases , Benzamidas , Línea Celular Tumoral , Cromosomas Humanos Par 9/genética , ADN de Neoplasias/genética , Inhibidores Enzimáticos/uso terapéutico , Amplificación de Genes , Humanos , Mesilato de Imatinib , Hibridación Fluorescente in Situ , Leucemia-Linfoma de Células T del Adulto/tratamiento farmacológico , Leucemia-Linfoma de Células T del Adulto/enzimología , Datos de Secuencia Molecular , Piperazinas/uso terapéutico , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/genética , Pirimidinas/uso terapéuticoRESUMEN
Classical t(11;14)(q13;q32) involving IGH-CCND1 is typically associated with aggressive CD5-positive mantle cell lymphoma (MCL). Recently, we identified the IGK variant of this translocation, t(2;11)(p11;q13), in three patients with a leukemic small-cell B-non-Hodgkin lymphoma. In all cases, rearrangements of the IGK and CCND1 genes were demonstrated by fluorescence in situ hybridization. Moreover, we mapped the 11q13 breakpoint of this variant translocation in the 3' region of CCND1 which contrasts with the 5' breakpoints in a standard t(11;14)(q13;q32). Expression of cyclin D1 was shown in two cases analyzed either at diagnosis or during disease progression. All three patients were asymptomatic at presentation and no initial therapy was required. One patient died of a progressive disease 58 months from diagnosis, and two patients showed stable disease after 12 months of follow-up. In two analyzed cases, mutated IGVH genes were identified. Our findings indicate that variant t(2;11)(p11;q13) does not typify a classical MCL but possibly a more indolent leukemic lymphoma originating from an antigen experienced (mutated) B cell.
Asunto(s)
Cromosomas Humanos Par 11/genética , Cromosomas Humanos Par 2/genética , Ciclina D1/genética , Reordenamiento Génico de Cadena Pesada de Linfocito B/genética , Inmunoglobulinas/genética , Linfoma no Hodgkin/genética , Translocación Genética/genética , Adulto , ADN de Neoplasias/análisis , Progresión de la Enfermedad , Femenino , Variación Genética , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/genética , Hibridación Fluorescente in Situ , Cariotipificación , Leucemia/genética , Leucemia/inmunología , Leucemia/patología , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/inmunología , Leucemia Linfocítica Crónica de Células B/patología , Linfoma de Células del Manto/genética , Linfoma de Células del Manto/inmunología , Linfoma de Células del Manto/patología , Linfoma no Hodgkin/inmunología , Linfoma no Hodgkin/patología , Masculino , Persona de Mediana Edad , Mutación , Recurrencia Local de Neoplasia/genética , Recurrencia Local de Neoplasia/patologíaRESUMEN
A patient with mantle cell lymphoma (MCL) of the pleomorphic blastoid subtype is reported. The disease was clinically aggressive and refractory to chemotherapy, and the patient survived only 2 months. Cytogenetically, a t(11;19;14)(q13;q13;q32) was found. Fluorescent in situ hybridization (FISH) and molecular analyses demonstrated involvement of the BCL1/CCND1 locus in a three-way translocation. In addition, subclonal abnormalities of the region 8q24 manifested either as a t(8;22)(q24;q11)/CMYC rearrangement or trisomy 8 were identified. The pathogenetic impact of this very uncommon association of BCL1/CCND1 and CMYC rearrangements in MCL is discussed and the literature is reviewed.
Asunto(s)
Ciclina D1/genética , Linfoma de Células del Manto/genética , Linfoma de Células del Manto/patología , Proteínas Proto-Oncogénicas c-myc/genética , Translocación Genética/genética , Trisomía/genética , Anciano , Cromosomas Humanos/genética , ADN de Neoplasias/genética , Femenino , Humanos , Inmunohistoquímica , Hibridación Fluorescente in Situ , Masculino , Persona de Mediana Edad , Resultado del TratamientoRESUMEN
Previous studies have shown that activating mutations of c-KIT/PDGFRA, potential therapeutic targets for imatinib mesylate, are implicated in the pathophysiology of gastrointestinal stromal tumours (GISTs). In this study, GISTs from 37 patients enrolled in an European Organisation for Research and Treatment of Cancer (EORTC) phase I/II clinical study of imatinib were examined for mutations of c-KIT/PDGFRA in order to explore whether the mutational status of the tumour predicts the clinical response to therapy. Mutations were screened by denaturing high-pressure liquid chromatography (DHPLC) and characterised by bi-directional DNA sequencing. Activating mutations of c-KIT or PDGFRA were found in 29 (78%) and 2 (6%) GISTs, respectively. Most c-KIT mutations involved exon 11 (n=24; 83%), all but one being an in-frame deletion; no isolated point mutations were found. The other c-KIT mutations included exon 9 AY 502-503 duplication (n=4; 14%) and exon 13 Lys-->Glu(642) missense mutation (n=1; 3%). Two tumours with no detectable c-KIT mutations demonstrated PDGFRA Asp-->Glu(842) amino acid substitutions. Patients with GISTs harbouring exon 11 mutations were more likely to achieve a partial response (PR) on imatinib therapy (83%) than all of the others (23%). The overall survival and progression-free survival rates for the entire group at 106 weeks were 78.3% and 46.9%, respectively. Based on a Kaplan-Meier analysis, patients with GISTs harbouring c-KIT mutations had longer median survival times and were less likely to progress than the other patients. These findings indicate that the mutational status of the c-KIT/PDGFRA oncoproteins could be useful to predict the clinical response of patients imatinib therapy.
Asunto(s)
Antineoplásicos/uso terapéutico , Neoplasias Gastrointestinales/tratamiento farmacológico , Mutación/genética , Piperazinas/uso terapéutico , Proteínas Proto-Oncogénicas c-kit/genética , Pirimidinas/uso terapéutico , Adulto , Anciano , Secuencia de Aminoácidos , Benzamidas , Análisis Mutacional de ADN , Femenino , Neoplasias Gastrointestinales/genética , Genotipo , Humanos , Mesilato de Imatinib , Masculino , Persona de Mediana Edad , Análisis de Supervivencia , Resultado del TratamientoRESUMEN
We here report the clinical, cytogenetic, fluorescence in situ hybridization (FISH), and Southern blot data on 14 patients with a myeloid malignancy and structural aberration of chromosome band 11q23 associated with overrepresentation or amplification of the MLL gene. The number of copies of MLL varied from three (two cases) to a cluster consisting of multiple hybridization spots. Together with previous reports, available data indicate that amplification of 11q23/MLL is a recurrent genetic change in myeloid malignancy. It affects mainly elderly patients and is often associated with dysplastic bone marrow changes or with complex karyotypic aberrations, suggestive of genotoxic exposure. It is associated with a poor prognosis. In addition, FISH analysis of nine cases with additional 11q probes showed that the overrepresented chromosomal region is generally not restricted to MLL, and Southern blot analysis indicated that amplification does not involve a rearranged copy of this gene. The significance of MLL amplification and the mechanisms by which it could play a role in leukemogenesis and/or disease progression remain to be elucidated.
Asunto(s)
Cromosomas Humanos Par 11/genética , Proteínas de Unión al ADN/genética , Amplificación de Genes/genética , Leucemia Mieloide/genética , Leucemia Mieloide/patología , Proto-Oncogenes , Factores de Transcripción , Adulto , Anciano , Anciano de 80 o más Años , Southern Blotting , Femenino , Dosificación de Gen , N-Metiltransferasa de Histona-Lisina , Humanos , Inmunofenotipificación , Hibridación Fluorescente in Situ , Cariotipificación , Leucemia Mieloide/diagnóstico , Masculino , Persona de Mediana Edad , Proteína de la Leucemia Mieloide-Linfoide , Estudios RetrospectivosRESUMEN
The genetic mechanisms underlying the genesis, disease progression, and high-grade transformation of marginal zone B-cell lymphoma (MZBCL) are poorly understood. We analyzed 33 cases of histologically and immunophenotypically well-characterized MZBCL (12 extranodal, 11 nodal, and 10 splenic MZBCL; 27 at primary diagnosis and six during the course of disease) by dual-color interphase fluorescence in situ hybridization (FISH) for deletions of tumor suppressor genes. We investigated loci known to play a role in the genesis or disease progression of other subtypes of lymphoid malignancies, namely the P53 gene (17p13), the retinoblastoma gene (RB, 13q14), the D13S25 locus (13q14), and the P16(INK4A) gene (9p21). Heterozygous deletions of P53 were detected in three out of the 33 cases, including two splenic and one extranodal MZBCL. One of these patients was analyzed at primary diagnosis and two during the course of disease. Heterozygous deletions of the RB gene (nodal MZBCL) and D13S25 (splenic MZBCL) were found in one case each. P16 deletions were not detected in any of our cases. We conclude that deletions of the analyzed tumor suppressor genes are relatively rare in MZBCL, which contrasts with the findings in some other subtypes of NHL.
Asunto(s)
Eliminación de Gen , Genes de Retinoblastoma/genética , Genes p16/genética , Genes p53/genética , Linfoma de Células B/genética , Anciano , Femenino , Humanos , Hibridación Fluorescente in Situ , Interfase , Masculino , Persona de Mediana EdadRESUMEN
Translocation t(12;21)(p13;q22) is the most frequent cytogenetic abnormality in childhood acute lymphoblastic leukemia (ALL) and is generally associated with favorable prognosis. In this report, we assessed the value of dual-color interphase fluorescence in situ hybridization (FISH) for the detection of t(12;21). Fifty-three patients were screened for ETV6/CBFA2 fusion by means of FISH, using two cosmid probes mapped on ETV6 and on CBFA2, respectively. The cut-off value (mean + three standard deviations) for positivity established on control patients was 9.3%. A comparison between FISH and molecular methods [reverse-transcriptase polymerase chain reaction/Southern blot (RT-PCR/SB)] was possible in 52 patients: 34 of 52 (65.4%) showed negative results with both approaches, and 13 of 52 (25%) were positive; 5 of 52 (9.6%) showed discrepancies: four patients who were positive using RT-PCR/SB were negative using FISH. Conversely, one patient negative when using RT-PCR/SB was positive with FISH. Further investigations on this patients, cytogenetically characterized by add(12p), showed an atypical breakpoint on ETV6, located 5' to the common breakpoint. Compared with RT-PCR and SB, dual-color interphase FISH with the cosmid probe set proved to be highly specific but showed limited sensitivity.
Asunto(s)
Cromosomas Humanos Par 12 , Cromosomas Humanos Par 21 , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Translocación Genética , Southern Blotting , Niño , Preescolar , Femenino , Humanos , Hibridación Fluorescente in Situ , Interfase , Recuento de Leucocitos , Masculino , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Reciprocal translocations involving the MLL gene on chromosome band 11q23 have been observed in both acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL). In AML, identification of MLL breakpoints is an important prognostic factor. Breakpoints are clustered in an 8 kb DNA fragment (bcr) and can be detected by Southern blotting or fluorescence in situ hybridization (FISH) analysis. Our objective in this study was to design a DNA probe set that enables optimal detection of MLL rearrangements using interphase FISH. Two PAC clones, 217A21 and 167K13, spanning the MLL gene with a minimal overlap in the bcr were isolated and labeled. Twenty-seven AML/ALL patients with cytogenetic 11q23 abnormalities, seven AML/ALL patients without 11q23 abnormalities but MLL rearrangement by Southern blotting, and eight healthy donors were analyzed by FISH. We compared this double-color FISH analysis with FISH using a YAC clone (yB22B2) and with Southern blotting. The PAC probe combination detects an MLL breakpoint in all cases with MLL rearrangement detected by Southern blotting except for cases with a partial tandem duplication detected by reverse transcriptase-polymerase chain reaction (RT-PCR). FISH using the PAC probes also detected MLL breakpoints in four cases with MLL deletions telomeric to the breakpoint that could not be detected by the single probe yB22B2. This new probe set provides a reliable and rapid assay for the diagnosis of AML and ALL patients with MLL/11q23 breakpoints.
Asunto(s)
Rotura Cromosómica/genética , Cromosomas Humanos Par 11/genética , Sondas de ADN/genética , Proteínas de Unión al ADN/genética , Hibridación Fluorescente in Situ/métodos , Interfase/genética , Leucemia/genética , Proto-Oncogenes , Factores de Transcripción , Enfermedad Aguda , Adolescente , Adulto , Anciano , Bacteriófago P1/genética , Preescolar , Clonación Molecular , Mapeo Contig , Femenino , N-Metiltransferasa de Histona-Lisina , Humanos , Cariotipificación , Masculino , Persona de Mediana Edad , Proteína de la Leucemia Mieloide-Linfoide , Translocación Genética/genéticaRESUMEN
The value of dual-color fluorescence in situ hybridization (FISH) for the detection of inv(16), using two contigs of cosmid probes mapping on both sides of the chromosome 16p breakpoint region, was evaluated in 23 acute myeloid leukemias (AML) in different phases of the disease. At diagnosis interphase FISH detected inv(16) in 19/19 (100%) cases with conventional cytogenetics (CC) evident aberration and excluded the rearrangement in two patients with CC suspected inv(16). Moreover, it also identified an associated del(16p) in two patients. At relapse, it revealed the inv(16) in 8/8 (100%) studied cases. These results were concordant with those of reverse transcriptase-polymerase chain reaction (RT-PCR). From 13 patients who obtained at least one complete remission (CR), 31 follow-up samples were analyzed using interphase FISH. Twenty-nine specimens scored negative for inv(16) and two were positive. RT-PCR detected CBFbeta/MYH11 transcripts in four of the nine CR samples analyzed, being more sensitive than interphase FISH. Eight of the 13 patients relapsed at a median time of 6.5 months (range 1-15) from the last negative FISH analysis. Of the two patients with positive FISH in CR, one relapsed soon after. At diagnosis and relapse, interphase-FISH proved to be an effective technique for detecting inv(16) appearing more sensitive than CC. Prospective studies with more frequent controls and possibly additional FISH probes are needed to assess the value of interphase FISH for minimal residual disease (MRD) and relapse prediction.
Asunto(s)
Inversión Cromosómica , Cromosomas Humanos Par 16/ultraestructura , Hibridación Fluorescente in Situ/métodos , Interfase , Leucemia Mieloide/genética , Enfermedad Aguda , Adolescente , Adulto , Anciano , Niño , Preescolar , Cromosomas Humanos Par 16/genética , Cósmidos/genética , Progresión de la Enfermedad , Femenino , Estudios de Seguimiento , Humanos , Cariotipificación , Leucemia Mieloide/epidemiología , Leucemia Mieloide/patología , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/genética , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Gastric low grade MALT lymphomas show a pattern of somatic mutations in their rearranged immunoglobulin genes, indicative of antigen selection. This provides evidence for antigen stimulation in the lymphomagenesis. Gastric diffuse large B cell lymphomas develop secondary to low grade MALT lymphoma or de novo. To study whether antigen-selection is also a feature of primary diffuse large B cell lymphomas, we analysed somatic mutations in the rearranged immunoglobulin heavy chain (IgH) variable genes (VH). The rearranged VH genes of six cases of gastric primary diffuse large B cell lymphoma were amplified from genomic or complementary DNA by a VH gene family-specific polymerase chain reaction method. The PCR products were directly sequenced and were compared to published germline sequences to analyse somatic mutations. Similarly to low grade MALT lymphomas 5/6 primary diffuse large B cell lymphomas show a pattern of somatic mutation in their rearranged VH genes, indicative of antigen selection and suggesting a role for antigens in lymphomagenesis. One case showed bi-allelic VH gene rearrangements, which were non-functional due to extensive deletions. Antigen selection could not be demonstrated or excluded. Antigen selection is a common feature in most analysed primary diffuse large B cell lymphomas, although some heterogeneity in the mechanisms involved in the lymphomagenesis of gastric primary diffuse large B cell lymphomas has not been excluded entirely (case 4).
Asunto(s)
Reordenamiento Génico , Cadenas Pesadas de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/genética , Linfoma de Células B/genética , Linfoma de Células B Grandes Difuso/genética , Neoplasias Gástricas/genética , Anciano , Anciano de 80 o más Años , Secuencia de Aminoácidos , Antígenos de Neoplasias/genética , Secuencia de Bases , Femenino , Humanos , Inmunofenotipificación , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Mutación , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
In order to gain insight into immunoglobulin (Ig) and T cell receptor (TCR) gene rearrangements in adult acute lymphoblastic leukemia (ALL), we studied 48 adult patients: 26 with precursor-B-ALL and 22 with T-ALL. Southern blotting (SB) with multiple DNA probes for the IGH, IGK, TCRB, TCRG, TCRD and TAL1 loci revealed rearrangement patterns largely comparable to pediatric ALL, but several differences were found for precursor-B-ALL patients. Firstly, adult patients showed a lower level of oligoclonality in the IGH gene locus (five out of 26 patients; 19%) despite a comparable incidence of IGH gene rearrangements (24 out of 26 patients; 92%). Secondly, all detected IGK gene deletions (n = 12) concerned rearrangements of the kappa deleting element (Kde) to Vkappa gene segments, which represent two-thirds of the Kde rearrangements in pediatric precursor-B-ALL and only half of the Kde rearrangements in mature B cell leukemias. Thirdly, a striking predominance of immature Ddelta2-Ddelta3 cross-lineage recombinations was observed (seven out of 16 TCRD rearrangements; 44%), whereas more mature Vdelta2-Ddelta3 gene rearrangements occurred less frequently (six out of 16 TCRD rearrangements; 38% vs >70% in pediatric precursor-B-ALL). Together these data suggest that the Ig/TCR genotype of precursor-B-ALL is more immature and more stable in adults than in children. We also evaluated whether heteroduplex analysis of polymerase chain reaction (PCR) products of rearranged Ig and TCR genes can be used for identification of molecular targets for minimal residual disease (MRD) detection. Using five of the major gene targets (IGH, IGK, TCRG, TCRD and TAL1 deletion), we compared the SB data and heteroduplex PCR results. High concordance between the two methods ranging from 96 to 100% was found for IGK, TCRG and TAL1 genes. The concordance was lower for IGH (70%) and TCRD genes (90%), which may be explained by incomplete or 'atypical' rearrangements or by translocations detectable only by SB. Finally, the heteroduplex PCR data indicate, that MRD monitoring is possible in almost 90% of adult precursor-B-ALL and >95% of adult T-ALL patients.
Asunto(s)
Envejecimiento/genética , Reordenamiento Génico de Linfocito T , Reordenamiento Génico , Genes de Inmunoglobulinas , Reacción en Cadena de la Polimerasa/métodos , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Proteínas Proto-Oncogénicas , Factores de Transcripción , Adolescente , Adulto , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Sondas de ADN , ADN de Neoplasias/genética , Proteínas de Unión al ADN/genética , Femenino , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas kappa de Inmunoglobulina/genética , Masculino , Persona de Mediana Edad , Neoplasia Residual , Sensibilidad y Especificidad , Proteína 1 de la Leucemia Linfocítica T AgudaRESUMEN
The inv(16)(p13q22) masked by different translocations was detected by fluorescence in situ hybridization (FISH) and confirmed by molecular analysis in three adult patients presenting with acute myeloid leukemia (AML)-M2 (cases 1 and 3) and M4Eo (case 2). Cytogenetic analysis revealed 47,XX,t(9;16)(p23;p13),+22 (case 1); 46,XX,t(1;16)(p32;p13) (case 2); and 46,XY,?del(16)(q22) (case 3). Using a panel of probes for chromosomes 1, 9, 16, and 20 as well as probes to detect inv(16), i.e., two cosmid contigs hybridizing proximally and distally to the 16p13 breakpoint, FISH demonstrated inv(16) involving the derivative 16 as well as reciprocal translocations between 16q22-qter and 9p24 (case 1), 1p32 (case 2), and 20q13 (case 3). In addition, a small interstitial del(16)(p13p13) proximal to the MYH11 breakpoint was detected in case 1. Reverse transcriptase-polymerase chain reaction (RT-PCR) and Southern blot analysis showed a CBFB-MYH11 fusion transcript and MYH11 rearrangement, respectively, in all three cases. We conclude that: 1) inv(16) can be masked by other structural abnormalities involving chromosome 16; 2) some of the so-called variant translocations not explored at the molecular level may in fact represent a masked inv(16); and 3) FISH, RT-PCR, and Southern blot analyses are reliable tools to detect masked inv(16) and should be applied in all AML cases with structural changes of chromosome 16.
Asunto(s)
Inversión Cromosómica , Cromosomas Humanos Par 16/genética , Hibridación Fluorescente in Situ/métodos , Leucemia Mieloide Aguda/genética , Translocación Genética/genética , Adulto , Southern Blotting , Cromosomas Humanos Par 1/genética , Cromosomas Humanos Par 20/genética , Cromosomas Humanos Par 9/genética , Femenino , Humanos , Cariotipificación , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mielomonocítica Aguda/diagnóstico , Leucemia Mielomonocítica Aguda/genética , Masculino , Persona de Mediana Edad , Proteínas de Fusión Oncogénica/genética , Reacción en Cadena de la PolimerasaRESUMEN
Abnormalities of chromosome 1q21 are common in B-cell malignancies and have been associated with a poor response to therapy. The nature of the involved gene(s) on chromosome 1q21 remains unknown. A cell line (CEMO-1) has recently been established from a patient with precursor-B-cell acute lymphoblastic leukemia (ALL), which exhibited a t(1;14)(q21;q32). To identify the gene involved in this translocation, we have cloned both rearranged IGHJ alleles using long-distance inverse polymerase chain reaction (LDI-PCR). Two IGHJ fragments were amplified from CEMO-1 DNA and sequenced. One allele showed novel sequences upstream of JH5 with no homology to either IGH or any other sequences on the databases. Using a single-copy Xho I fragment immediately 5' of JH5, PAC clones were isolated and mapped to chromosome 1q21 on normal metaphases by fluorescence in situ hybridization (FISH), confirming that this allele represented the t(1;14)(q21;q32) breakpoint. Sequence analysis of the 1q21 Xho I fragment showed identity with an expressed sequence tag (EST), and this probe was therefore used to probe Northern blots. Two transcripts of 6.3 kb and 4.2 kb expressed at low level in mRNA from all tissues were detected: a third transcript of 1.6 kb was expressed only in thymus, spleen, and small intestine. Full-length BCL9 cDNA clones were obtained from a normal human fetal brain cDNA library supplemented by 5' and 3' RACE. Sequence analysis predicted a protein of 1394 amino acids containing 18% proline, 11% glycine, 11% serine, and 6% methionine, but no recognizable protein motifs or significant homologies to any other known proteins. The CEMO-1 1q21 breakpoint fell within the 3' UTR of the BCL9 gene. Low-level expression of BCL9 was detected in Epstein-Barr virus-transformed normal B cells by Northern blot; in contrast, abundant BCL9 expression was observed in CEMO-1, indicating that deregulated expression of this gene was one pathological consequence of the translocation. Screening of a panel of 39 B-cell malignancies with 1q abnormalities by Southern blot showed one additional case with a breakpoint in the 3' UTR of BCL9, indicating that this was a recurrent breakpoint. FISH analysis using an 850-kb YAC spanning BCL9 identified a further case with t(1;22)(q21;q11) causing juxtaposition of BCL9 to the IGlambda locus. Other breakpoints were heterogeneous, falling both centromeric (10 cases) and telomeric (10 cases) of the BCL9 gene. These data suggest that BCL9 may be the target of translocation in some B-cell malignancies with abnormalities of 1q21 and that deregulated BCL9 expression may be important in their pathogenesis.
Asunto(s)
Cromosomas Humanos Par 14/ultraestructura , Cromosomas Humanos Par 1/genética , Cromosomas Humanos Par 1/ultraestructura , Genes , Proteínas de Neoplasias/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Translocación Genética , Adulto , Secuencia de Aminoácidos , Secuencia de Bases , Northern Blotting , Línea Celular Transformada , Mapeo Cromosómico , Clonación Molecular , ADN de Neoplasias/genética , Regulación Leucémica de la Expresión Génica , Humanos , Masculino , Datos de Secuencia Molecular , Proteínas de Neoplasias/biosíntesis , Especificidad de Órganos , Leucemia-Linfoma Linfoblástico de Células Precursoras B/metabolismo , ARN Mensajero/biosíntesis , ARN Mensajero/genética , ARN Neoplásico/biosíntesis , ARN Neoplásico/genética , Factores de TranscripciónRESUMEN
In a case of follicular center cell lymphoma (FCCL) without evidence of histologic progression towards a high-grade lymphoma, t(9;22)(q34;q11) was found simultaneously with a t(14;18)(q32;q21) and a t(8;14)(q24;q32). Molecular studies of this case showed BCL2 and MYC rearrangements in addition to the rearrangements of immunoglobulin heavy (IGH) and lambda (IGL) loci. Investigation of the t(9;22) using Southern blot and RT-PCR analysis failed to detect M-bcr or m-bcr rearrangements of BCR. Two-color fluorescence in situ hybridization (FISH) with ABL and BCR probes revealed presence of a "fusion" signal, but its atypical localization [der(9)] and gene order [cen-ABL-BCR-tel] indicated that this translocation differed from the t(9;22) in chronic myeloid leukemia and did not involve either ABL or BCR. In addition, further FISH analysis using 9q34- and 22q11-specific probes localized the breakpoint on chromosome 9 distal to the NOTCH1 gene and the breakpoint on 22q11 in the IGL gene cluster. These results indicate an IGL-mediated rearrangement of an unknown gene at 9q34 that together with BCL2 and MYC might be involved in the lymphomagenesis of the present case of FCCL and perhaps in other cases of non-Hodgkin lymphoma in which t(9;22) is sporadically occurring.
Asunto(s)
Cromosomas Humanos Par 22/genética , Cromosomas Humanos Par 9/genética , Reordenamiento Génico , Linfoma Folicular/genética , Translocación Genética/genética , Southern Blotting , Bandeo Cromosómico , Genes abl , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas lambda de Inmunoglobulina/genética , Hibridación Fluorescente in Situ , Cariotipificación , Linfoma Folicular/patología , Masculino , Persona de Mediana Edad , Familia de Multigenes , Reacción en Cadena de la PolimerasaRESUMEN
Clinical, cytogenetic, fluorescence in situ hybridization (FISH), and Southern blot data of 18 patients with different subtypes of B-cell non-Hodgkin's lymphoma, cytogenetically characterized by partial trisomy 12, are presented. These chromosomal changes occurred predominantly in clinically progressive chronic lymphocytic leukemia, mixed cell type, and advanced-stage follicle center cell lymphoma at the time of relapse or transformation into diffuse large cell lymphoma. Partial trisomy 12 consistently included the long arm of chromosome 12, either completely or partially, and resulted from dup(12q) or other rearrangements involving chromosome 12. The duplications were cytogenetically identified as dup(12)(q13q23), dup(12)(q13q22), or dup(12)(q13q15) in follicle center cell lymphoma or t(14;18)-positive diffuse large cell lymphoma; dup(12)(q13q22) or dup(12)(q13q24) in chronic lymphocytic leukemia; and dup(12)(q13q21) in a case of t(14;18)-negative diffuse large cell lymphoma. FISH, using library probes and a panel of YAC probes, mapped along the long arm of chromosome 12, confirmed the cytogenetic results in all cases analyzed except for three cases of t(14;18)-positive follicle center lymphoma or diffuse large cell lymphoma with dup(12q). In these cases, FISH showed similar, possibly identical, duplications, which involved a region more centromeric (12q11-21) than assumed by karyotypic analysis (12q13-22 or 12q13-23) and included alphoid DNA sequences, a combination hitherto unknown. In addition, commonly duplicated regions of chromosome 12 could be defined: 12q11-21, including alphoid DNA sequences for follicle center cell lymphoma or t(14;18)-positive diffuse large cell lymphoma, 12q13-22 for chronic lymphocytic leukemia, and 12p13-q15 for marginal zone cell lymphoma, all of which overlapped in 12q13-15. Whether these regions, especially 12q13-15, may contain genes which are important in malignant transformation or disease progression of B-cell lymphoproliferative malignancies characterized by complete or partial trisomy 12 remains to be determined.