Leucemia linfoide crónica: aspectos citogenéticos y moleculares / Chronic Lymphoid Leukemia: cytogenetic and molecular aspects

Introducción: La leucemia linfoide crónica es un trastorno linfoproliferativo caracterizado por la acumulación de linfocitos pequeños de aspecto maduro en sangre periférica, médula ósea y tejidos linfoides con un período de vida prolongado. Presenta una gran variabilidad clínica y genética. Objetivo: Describir los aspectos citogenéticos y moleculares de la leucemia linfoide crónica. Métodos: Se realizó revisión de la literatura en inglés y español, a través del sitio web PubMed y el motor de búsqueda Google académico, de artículos publicados en los últimos 5 años. Se hizo un análisis y resumen de la bibliografía revisada. Desarrollo: En la leucemia linfoide crónica están presentes alteraciones citogenéticas frecuentes como la deleción de los cromosomas 13q, 11q y 17p, así como la trisomía 12, que unido al conocimiento del estado mutacional del gen de la región variable de la cadena pesada de la inmunoglobulina, y otras mutaciones somáticas en diferentes genes, así como a variables clínicas y de laboratorio permiten la estratificación pronóstica de los pacientes. Conclusiones: El diagnóstico a través de los estudios citogenéticos convencionales estimulados con mitógenos, la hibridación in situ por fluorescencia y la secuenciación génica permite una mayor comprensión de la biología de la enfermedad, así como tomar decisiones terapéuticas más personalizadas(AU)

Introduction: Chronic B lymphoid leukemia is a lymphoproliferative disorder characterized by the accumulation of small, mature-looking lymphocytes in peripheral blood, bone marrow and lymphoid tissues with a long life span. It has great clinical and genetic variability. Objective: To describe the cytogenetic and molecular aspects of the disease. Methods: A review of the literature in English and in Spanish was carried out, in the PubMed website and using the search engine of Google Scholar, for articles published in the last five years. We performed analysis and summary of the reviewed bibliography. Development: In chronic lymphoid leukemia, frequent cytogenetic alterations are present such as deletion of chromosomes 13q, 11q and 17p, as well as trisomy 12, which together with the knowledge of the mutational status of the gene for the variable region of the immunoglobulin heavy chain and other somatic mutations in different genes, as well as clinical and laboratory variables allows prognostic stratification of patients. Conclusions: Diagnosis through conventional mitogen-stimulated cytogenetic studies, fluorescence in situ hybridization and gene sequencing allow a better understanding of the biology of the disease, as well as making more personalized therapeutic decisions(AU)

Leucemia linfoide crónica de células B: revisión de sus aspectos etiopatogénicos, moleculares y pronósticos / B-cell chronic lymphoid leukemia: a review of its etiopathogenic, molecular and prognostic aspects

La leucemia linfoide crónica (LLC) es una neoplasia maligna que afecta principalmente a pacientes de mediana edad y ancianos. Se caracteriza por la proliferación de linfocitos morfológicamente maduros pero inmunoincompetentes que se acumulan en sangre periférica, médula ósea y tejido linfático. Presenta gran heterogeneidad clínica. Se describen diversos fenotipos, aunque predomina la expansión clonal de células B CD5+CD23+. Los factores pronósticos en la LLC incluyen el subgrupo citogenético, estado mutacional de inmunoglobulina, la expresión de ZAP-70, CD38 y CD49d. El tratamiento se basa en usar modernos algoritmos terapéuticos aprobados, que produzcan mayores respuestas y menores eventos secundarios, en lograr la remisión clínica completa y mejorar la calidad de vida de estos pacientes(AU)

Chronic lymphocytic leukemia (CLL) is a malignancy that mainly affects middle-aged and elderly patients. It is characterized by the proliferation of morphologically mature but immunoincompetent lymphocytes that accumulate in blood, bone marrow and lymphatic tissue. It presents great clinical heterogeneity. Several phenotypes are described, although the clonal expansion of CD5 + CD23 + B cells predominates. Prognostic factors include the cytogenetic subgroup, immunoglobulin mutational status, expression of ZAP-70, CD38, and CD49d. The treatment is based on using modern approved therapeutic algorithms that produce greater responses and minor secondary events, to achieve complete clinical remission and to improve the quality of life of these patients(AU)

Classification of Acute Leukemias – Past, Present,and Future.

The perspective of the classification of any disease is to treat them according to their biologic behavior. Standard criteria to distinguish between myeloid and lymphoid acute leukemias were laid down as the first of its kind, by the French-American-British (FAB) working group. The FAB classification had a cursory correlation with clinical outcome, poor concordance owing to inter-observer variation, and failure to incorporate cytogenetic data. Hence, the World Health Organization (WHO) classification of leukemias evolved in 1997 with the goal of improving the objectivity and reproducibility, which had incorporated cytogenetic abnormalities and immunology as principal designating criteria, other than the morphology. Major changes were made in the subsequent editions of WHO classification, incorporating newer genetic abnormalities such as mutations of nucleophosmin member 1, CCAAT/enhancer-binding protein alpha; renaming of the existing classes, etc. The role of the genes encoding guanine nucleotide-binding protein gamma 11, amphiregulin, and ceruloplasmin; the biomarkers platelet factor 4 and connective tissue-activating peptide III, complement fragment C3a; the mRNA coding for plexin C1, leukotriene B4 receptor 1, and Immunoglobulin superfamily member 2; mixed lineage leukemia gene rearrangement in the prognosis of leukemias is proven. Thus, the approach of diagnostics using cytogenetics and immunophenotyping may further be modified.

Epidemiologia da leucemia linfocítica crônica e leucemia linfocítica crônica familiar / Epidemiology of chronic lymphocytic leukemia and familial chronic lymphocytic leukemia

A leucemia linfocítica crônica (LLC) é uma doença linfoproliferativa crônica (DLPC) que apresenta características epidemiológicas peculiares; acomete indivíduos com idade mais avançada (não ocorre em crianças e é rara abaixo dos 30 anos) e a sua incidência varia conforme a origem étnica dos pacientes. É a leucemia mais freqüente nos países ocidentais enquanto muito rara nos orientais. A sua etiologia não está ainda esclarecida, não se conhecendo fatores ambientais que mostrem forte associação com o surgimento da doença. Não tem relação com radiação ionizante (sobreviventes da bomba atômica não apresentaram aumento na incidência da LLC), assim como não foi demonstrada associação com agentes tóxicos ou virais específicos. A LLC familiar é assim denominada quando, pelo menos, dois membros de uma família apresentam LLC e o diagnóstico nestes pacientes costuma ocorrer em idade mais precoce. A ocorrência é maior em parentes de primeiro grau (irmãos, filhos), podendo, porém, afetar parentes mais distantes. Os familiares de pacientes com LLC apresentam maior freqüência de outras DLPC (risco relativo 30x maior) e de proliferação monoclonal de linfócitos B (13 por cento-18 por cento) e parece que o HLADR1.11 está implicado na LLC familiar, pelo menos em algumas populações.

Chronic lymphocytic leukemia (CLL) is a chronic lympho-proliferative desorder (CLPD) with peculiar epidemiologic characteristics. It is a disease of the elderly, which is very rare in under 30-year-old individuals and absent among children. Its incidence largely varies according to the ethnical origin: CLL is the most common leukemia in Western countries while it is rarely seen in Eastern countries. The etiology of CLL is still unknown. Environmental factors such as exposure to ionizing radiation (atomic bomb survivors did not show an increased incidence of CLL) or toxic or viral agents are not associated to the occurrence of CLL. Familiar CLL is characterized when at least two members of the same family develop CLL The diagnosis of familiar CLL seems to be made 10 to 20 years earlier than sporadic cases. CLL occurs more frequently among siblings and offspring although it may appear in other members such as cousins. The frequency of other CLPD (relative risk is 30 times greater) and monoclonal CD5+B cells proliferations (13.5 percent-18 percent) are higher among "healthy" members of familial CLL and HLADR1/11 is implicated in at least some populations with familial CLL.

Rapid identification of single nucleotide polymorphisms by fluorescence-based capillary electrophoresis

We describe the application of two different fluorescence-based techniques (ddNTP primer extension and single-strand conformation polymorphism (SSCP)) to the detection of single nucleotide polymorphisms (SNPs) by capillary electrophoresis. The ddNTP primer extension technique is based on the extension, in the presence of fluorescence-labeled dideoxy nucleotides (ddNTP, terminators), of an unlabeled oligonucleotide primer that binds to the complementary template immediately adjacent to the mutant nucleotide position. Given that there are no unlabeled dNTPs, a single ddNTP is added to its 3' end, resulting in a fluorescence-labeled primer extension product which is readily separated by capillary electrophoresis. On the other hand, the non-radioisotopic version of SSCP established in this study uses fluorescent dye to label the PCR products, which are also analyzed by capillary electrophoresis. These procedures were used to identify a well-defined SNP in exon 7 of the human p53 gene in DNA samples isolated from two human cell lines (CEM and THP-1 cells). The results revealed a heterozygous single-base transition (G to A) at nucleotide position 14071 in CEM cells, proving that both fluorescence-based ddNTP primer extension and SSCP are rapid, simple, robust, specific and with no ambiguity in interpretation for the detection of well-defined SNPs

Hipocomplementemia familiar en la leucemia linfoide crónica / Family hypocomplementaemia in chronic lymphocytic leukemia

Se efectuó la medición de la actividad de la vía clásica, alterna, factor B, factor D, así como la cuantificación de los componentes C1q, C3 y C4 del complemento en 31 pacientes con el diagnóstico de leucemia linfoide crónica (LLC) estadificados en 2 grupos: 14 pacientes en fase poco avanzada de la enfermedad (estadios 0, I, II) y 17 en fase avanzada (estadios III, IV); además en familiares sanos con primer grado de consanguinidad (PGC). Se demostraron deficiencias del complemento en el 70,5 porciento (12/17) de los pacientes en fase avanzada de la enfermedad, fundamentalmente una disminución de la actividad de la vía clásica y los niveles de C1q y C4. En los pacientes en fase poco avanzada se observaron deficiencias sólo en 3 (21,4 porciento). El 65 porciento de los familiares con PGC de los enfermos con LLC con alteraciones del complemento mostraron también deficiencias, lo que sugiere que en algunos enfermos con LLC la hipocomplementemia puede ser el reflejo de cierta predisposición genética

Estudio de los haplotipos HLA en leucemias / Study of HLA haplotypes in leukemias

Se determinaron las combinaciones alélicas de los genes HLA A y B en 144 pacientes con leucemias; 89 con leucemia linfoide aguda (LLA), 28 con leucemia mieloide aguda (LMA) y 27 con leucemia mieloide crónica (LMC), estraficados fenotípicamente en blancos, negros y mestizos, para evaluar la posible asociación entre los haplotipos HLA y esas enfermedades. Se utilizaron como controles 276 personas aparentemente estratificados también por el color de la piel. Los haplotipos A2-B12, A9-B12 y A2-B35 fueron los más frecuentes en el grupo total de enfermos y en los individuos blancos con LLA. La combinación de las especificidades A2 y B5 mostró un desequilibrio de asociación en el conjunto de enfermos y en los sujetos blancos con LMA

Chromosome fragility in hematologic disease presumably induced by environmental pollutants

Chromosome instability consists of chromosome abnoprmalities as multiple breaks in in metaphase chromosomes in syndromes of chromosaome instability such as fanconi's anemia (FA) which is mainly characaterized by bone marrow aplasia; some cases progress to acute leukemia. FA is a hereditary disease with recessive and monogenic transmission. This pair of mutated genes is related to chromosome fragility and therefore is responsible for the inefficiency of DNA repair. In normal individuals, these genes are assumed to be expressed normally, but their products ara insufficient to perform DNA repair when the intensity of the polluting agent lelads to exposure above basal levels. In the present study, the bone marrow of four patients with different hematologic diseases was submitted to cytogenetic analysis. Two had aplastic anemia, and one bad lymphoblastic leukemia. These patients were from towns unb the Amazon Region where the mercuri used for gold prospecting and the substances used and/or released in aluminium mining have been introduced into the environment. The last patient had fanconi's anemia and was used as a model in the discussion of the results. The cytogenetic findings were similar for all patients, the major ones being chromosome fragmentation and pulverization. In view of these findings, we believe that chromosome fragility, observed in the first three patients, presumably was induced by environmental pllutants

Detecçäo de mRNAs quiméricos BCR/ABL em pacientes portadores de leucemia mielóide crônica, leucemia linfóide aguda/mielóide aguda pela reaçäo de polimerizaçäo em cadeia (PCR) / Detection of BCR/ABL chimeric mRNAs in patients with chronic myeloid leukemia, acute lymphocytic leukemia/ acute myeloid by polimerase chain reaction (PCR)

A leucemia mielóide crônica (LMC) é uma doença caracterizada pela proliferaçao e acumulaçao de células mielóides e seus precursores, induzida pela transformaçao neoplásica de uma célula hematopoética pluripotente, a stem-cell. Esse clone possui uma anormalidade citogenética que se caracteriza pela translocaçao recíproca entre os cromossomos 9 e 22 [t(9;22) (q34;q11)], resultando na formaçao do cromossomo Philadelphia (Ph1). O resultado dessa translocaçao é a justaposiçao do gene BCR com o protooncogene ABL, originando um gene híbrido que codifica uma proteína anormal com atividade tirosina quinase exacerbada. A reaçao de polimerizaçao em cadeia (PCR) baseada na amplificaçao do cDNA híbrido BCR/ABL tem sido considerada um meio altamente sensível e específico para detecçao dessa patologia, em nível molecular. O RNA total foi extraído pelo método de isocianato de guanidina, de leucócitos de sangue periférico e (ou) medula óssea. A primeira fita (cDNA) foi sintetizada utilizando primer complementar à regiao 3' do mRNA quimérico BCR/ABL, transcriptase reversa (Super Script II - Gibco-BRL) segundo sugestao do fabricante. A reaçao de PCR foi realizada em duas etapas: a primeira utilizando primers complementares à regiao BCR e ABL, num total de 25 ciclos com temperatura de 94 graus Celsius, 49 graus Celsius e 72 graus Celsius para desnaturaçao, anelamento e extensao, respectivamente; na segunda etapa foram utilizados primers internos aos primeiros (Nested-PCR) num total de 35 ciclos com temperatura de anelamento de 60 graus Celsius (primers sintetizados pela Genomic - Engenharia Molecular, SP). O controle de todo o procedimento técnico foi realizado pela amplificaçao de uma regiao do gene ABL. A presença de bandas características da LMC foram visualizadas após eletroforese em gel de poliacrilamida 6 por cento e coloraçao por brometo de etídio. O método foi considerado extremamente sensível e rápido para a detecçao da t(9;22) quando comparada com a citogenética convencional.

Estudo cromossômico em leucemia, linfoma e câncer de pulmäo / Leukemias, lymphomas and lung cancer: chromosomic studies

Os autores descrevem os resultados da análise citogenética realizada em 16 pacientes portadores de leucemia, linfoma e câncer de pulmäo. Neste grupo forma observadas alteraçöes cromossômicas constitucionais, como duplicaçäo e translocaçäo, quebras cromossômicas preferenciais e variantes de heterocromatina constitutiva. Embora näo tenha sido possível detectar uma associaçäo direta destas características com a condiçäo neoplásica, taus achados em cerca de 37 porcento da amostra sugerem que algumas tenham desempenhado um papel no desenvolvimento maligno de seus portadores.

Alteraciones cromosomicas en leucémia / Chromosome aberrations in leukemia

Los progresos técnicos en la metodología para obtener cromosomas de células leucémicas han permitido lograr un mejor conocimiento de las alteraciones numéricas y/o estructurales. Dichos hallazgos citogenéticos son utilizados para clasificar distintos subgrupos de leucemias agudas y crónicas, pronosticar su evolución clínica y lograr un diagnóstico correcto en la elección del tratamiento más apropiado