Comprehensive genetic characterization of CLL: a study on 506 cases analysed with chromosome banding analysis, interphase FISH, IgVH status and immunophenotyping

Abstract

In CLL data from chromosome banding analysis (CBA) have been scarce due to the low proliferative activity of CLL cells in vitro. We improved the cultivation technique using an immunostimulatory CpG-oligonucleotide DSP30 and IL-2. A total of 506 CLL samples were analysed with CBA and interphase FISH using probes for the detection of trisomy 12, IgH rearrangements and deletions of 6q21, 11q22.3 (ATM), 13q14 (D13S25 and D13S319) and 17p13 (TP53). A total of 500 of 506 (98.8%) cases were successfully stimulated for metaphase generation and are subject to this study. Aberrations were detected in 415 of 500 (83.0%) cases by CBA and in 392 of 500 (78.4%) cases by FISH. CBA detected 832 abnormalities and FISH only 502. Therefore, CBA offers important information in addition to FISH. (1) CLL is characterized mainly by genomic imbalances and reciprocal translocations are rare. (2) A subgroup with complex aberrant karyotype (16.4%) is identified which is associated with an unmutated IgV H status and CD38 expression (P ¼ 0.034 and 0.02, respectively). (3) Additional abnormalities are detectable providing new biological insights into different CLL subclasses revealing a much more heterogeneous pattern of cytogenetic abnormalities as assumed so far based on FISH data only. Therefore, prospective clinical trials should evaluate the prognostic impact of newly available CBA data.

Stimulation of B-cell lymphoproliferations with CpG-oligonucleotide DSP30 plus IL-2 is more effective than with TPA to detect clonal abnormalities

“…Stimulation with DSP30 þ IL-2 allowed the detection of more abnormalities, more abnormal subclones and more complex karyotypes in CLL and in the majority of B-cell lymphoid neoplasms. As in earlier studies, 3,5,6 we think that CC is complementary to FISH analysis. Though FISH exploration using a large probe panel has yielded valuable results in lymphoproliferative diseases, it underestimates the heterogeneity of chromosomal aberrations.…”

“…These data are in agreement with the preliminary study of Dicker et al 3 concerning 132 CLL stimulated with DSP30 þ IL-2 alone. In a more recent study with a larger analyzed pool (n ¼ 506), 5 98.8% success was observed using CC with 83% aberrations. Our data revealed differences depending on the mitogene used: for both CLL and B-cell lymphoid malignancies, the number of metaphases per slide was higher with TPA stimulation, with a gain of 20 metaphases on the average (Po0.01).…”

“…The use of the immunostimulatory CpGoligonucleotide DSP30 to effectively induce cell cycle progression of CLL cells in vitro has been reported. [3][4][5][6] This proliferation is markedly enhanced upon the addition of interleukin-2 to cultures. 4,7 Recently, Haferlach et al 5 characterized 506 CLL stimulated with DSP30 þ IL-2.…”

“…Complexity of chromosomal changes, recently associated with unfavorable outcomes, can only be assessed with CC. 5,6 One important benefit of karyotype is that the prognostic subgroups defined by interphase FISH may be further subdivided if chromosomal rearrangement status is included. For instance, in our cohort, 23 CLL would have been assigned to the prognostically most-favorable group because of a 13q deletion as the sole abnormality detected by FISH, but six of them (10%) presented additional abnormalities on karyotypes, potentially changing their prognosis.…”

Chronic lymphocytic leukemia and treatment resistance in cancer: The role of the p53 pathway

“…27 We assessed the genetic complexity in relation to 17p-and TP53 mutation status in a cohort of 60 patients by the use of array-CGH and SNP-array analysis (500 K). 28 We separated three groups based on presence or absence of 17p-, TP53 mutation and none of these changes.…”