Study On Molecular Cytogenetic Abnormalities In B-lineage Lymphoid Malignancies

Objective:This study was aimed to clarify the molecular cytogenetic characterization in a case of B-lineage acute lymphoblastic leukemia (B-ALL) with t(14;14)(q11;q32) and trisomy 4, to explore the relations between immunophenotypic subtypes and prognosis in patients with diffuse large B-cell lymphoma (DLBCL), to establish the technique of interphase fluorescence in situ hybridization (FISH) in routine paraffin-embedded tissue and to investigate the incidence of common genetic abnormalities in Chinese patients with DLBCL and their relationships with clinical outcome.Methods:In the first part, we performed comprehensive cytogenetic and molecular cytogenetic analyses on a case of B-ALL with t(14;14)(q11;q32) and trisomy 4. Conventional RHG banding technique was employed for karyotypic analysis. Chromosome painting was carried out using whole chromosome paint (WCP) probe for chromosome 4. Two BAC clones flanking CEBPE locus at 14q11 were labeled by nick translation with biotin-16-dUTP and digoxigenin-11-dUTP, respectively. FISH analyses were performed on interphase and metaphase cells using dual-color break-apart IGH probe and 2 BAC probes flanking CEBPE locus, respectively and the multiprobe ALL panel.In the second part, immunohistochemical stain was used to evaluate the expression of CD10, BCL6 and MUM1 in 59 cases of DLBCL. The cases were then categorized into germinal center B-cell (GCB) and non-GCB subtypes. The overall survival rate was compared between the GCB and non-GCB groups. Interphase FISH analyses on paraffin-embedded tissue sections were performed to detect translocation and copy number increase of BCL6,BCL2 and c-MYC genes in 59 cases of DLBCL, using dual-color break-apart IGH probe, dual-color dual-fusion IGH/BCL2 probe and dual-color break-apart c-MYC probe, respectively. The relations between these genetic abnormalities and immunophenotypic subtypes or clinical outcome were explored. Results:1. Conventional RHG banding analysis revealed a karyotype of 47, XX, +4, t(14;14)(q11;q32), which was further confirmed with WCP and FISH. FISH analyses showed that translocation (14;14)(q11;q32) in B-ALL simultaneous involved the IGH and CEBPE genes. In B-ALL patients, the t(14;14)(q11;q32) may be associated with a better prognosis.2. The percentage of GCB subtype and non-GCB subtype in DLBCL were 29% and 71%,respectively. BCL6 expression and GCB phenotype could predict better overall survival (OS).3. The rates of BCL6 gene translocation and copy number increase in DLBCL were 24.1% and 17.2%, respectively. No relationship was found between BCL6 gene translocation or copy number increase and BCL6 expression, immunophenotypic subtypes or prognosis.4. The rates of BCL2 gene translocation and copy number increase in DLBCL were 1.7% and 22.2% respectively. BCL2 gene copy number increase was more common in non-GCB type (29%) than in GCB type (6%), but it was not related to BCL2 expression and prognosis.5. The rates of c-MYC gene translocation and copy number increase in DLBCL were 5.3 % and 19.3 % respectively. c-MYC translocation failed to correlate with immunophenotypic subtypes, but could predict worse outcome. Whereas c-MYC copy number increase was not related to molecular subtypes and prognosis.6. c-MYC gene copy number increase showed positive relationship with BCL6 and BCL2 copy number increase.Conclusions:1. Translocation (14;14)(q11;q32) simultaneously involving IGH and CEBPE genes in B-ALL is rare but recurrent genetic abnormality. It could identify a new subgroup. In B-ALL patients, t(14;14)(q11;q32) could predict better outcome.2. DLBCL can be subclassified into two distinct immunophenotypic subtypes: GCB-DLBCL and non-GCB-DLBCL. The GCB subtype of DLBCL is less frequent in our country. GCB subtype of DLBCL cases show better prognosis than non-GCB subtype of DLBCL cases.3. FISH analysis on paraffin-embedded tissue section is feasible and useful. 4. The rate of BCL6 gene translocation or copy number increase is high in DLBCL, and the detection of BCL6 gene translocation and copy number increase may help in the diagnosis and differential diagnosis.5. The occurrence of IHG/BCL2 translocation in DLBCL is significantly less in Chinese than Western populations. BCL2 gene copy number increase may play a crucial role in the pathogenesis of non-GCB subtype of DLBCL.6. In DLBCL patients, c-MYC translocation may be associated with a worse prognosis.