ALBERT

Description: Since January 2004, the French Ministry of Health supports a multicentric study aimed at detecting biological prognostic markers in B-CLL at Binet stage A, able to identify those patients in early clinical disease stage who will progress to a more advanced disease requiring early therapy. These include: 1/ detection by FISH of prognostically relevant cytogenetic abnormalities [trisomy 12, deletions of the ATM and P53 genes, and deletions at chromosomal band 13q14], 2/ cytoplasmic expression of ZAP-70 and CD38 by flow cytometry, 3/ IgVH gene analysis, and 4/ serum concentration levels of sCD23 and thymidine kinase. 550 patients from 10 institutions are currently enrolled in this study, 331 men and 219 women, with a median age at the time of diagnosis of 66 years (ranged from 26 to 100). To date interphase cytogenetic analyses by FISH have been carried out in leukemic cells of 484 patients using commercially available probes. For deletions, results are scored as A, B or C depending on the % of deleted lymphocytes [A = 20–100)]. The distribution of the prognostically relevant cytogenetic abnormalities is as follows: 63.6 % (n = 308) 13q14 deletions, 12.6% (n = 61) trisomies 12, 11.1% (n = 54) ATM gene deletions, and 16.6 % (n = 81) P53 gene deletions, all equally shared between men and women subgroups. Conventional cytogenetics (CC) have been performed in 391 out of the 484 patients. Results of the two techniques (FISH/CC) are highly discordant for detection of 13q-, 11q-, and 17p- deletions, respectively observed in 8.8 %, 3.6%, 4.4% of our cases by CC, and quite similar for detection of trisomy 12 (9.8% by CC). Nevertheless, FISH/CC discordance disappears for 11q- and 17p- detections when only FISH results from the C group are taken into account (5.7%, 4.3% deleted cells, respectively, in C group), showing the power of FISH to detect minor 11q- and 17p- clones in B-CLL at Binet stage A. In addition, CC detect abnormal karyotypes in 38.6 % of cases, including recurrent numerical abnormalities [21% (n=32) involving chromosome X, 3.3% chromosome 19], and structural rearrangements (5.3% at 14q32, and 3.9% 6q deletion). 15.9% of abnormal karyotypes (n = 24) are complex. IgVH gene analyses have been performed in 301 patients. As expected, adverse prognostic genetic markers [ATM and P53 deletions, trisomy 12] are significantly more frequent in cases with unmutated IgVH sequences (n = 83) as compared to mutated cases (n = 218) [χ 2 tests: P 〈 0.0001, P = 0.008, and P = 0.0004, respectively]. On the contrary, the prognostically favorable 13q14 deletion is more frequent in cases with mutated IgVH status [χ 2 test: P = 0.0002]. Comparison with the ZAP-70 status, analyzed by flow cytometry, shows less clear distributions of ATM and P53 deletions, trisomy 12 and 13q deletion, between the ZAP-70 + (n = 112) and ZAP-70 - (n=209) subgroups [χ 2 tests: P 〈 0.008, P = 0.32, P = 0.046, and P = 0.048, respectively]. By May 2006, this ongoing study designed for 700 patients should provide very helpful data on laboratory parameters important in the management of untreated B-CLL at Binet stage A.

Description: Background: Waldenstrom’s Macroglobulinemia(WM) is a rare B-cell disorder characterized by the accumulation of monoclonal lymphoplasmocytic cells and a serum monoclonal IgM protein. The genetic basis of this disorder is poorly understood. Molecular cytogenetic abnormalities differ from those reported for multiple myeloma, lymphomas, and B-cell chronic lymphocytic leukemia. One recurrent abnormality, deletion of the long arm of chromosome 6, was reported with a high prevalence (23–50%), but seems associated with advanced disease and clonal evolutions. Aims : As data on molecular changes in WM are limited, we used interphase fluorescence in situ hydidization (IP-FISH) to evaluate the prevalence of trisomy 4 in WM patients. Material and methods: 39 patients with a diagnosis of WM , according to the Workshop classification, were included: 28 men and 11 women. Mean age at diagnosis was 66 years (48–84 years). Mean percentage of bone marrow lymphoplasmocytic cells was 47% (13–97%). Conventional cytogenetic was performed after 72 or 96 hours culture with TPA, at diagnosis for 23 patients and 3.8 years median (6 months–9 years) after for 13. FISH study was carried on cytogenetic cryopreserved pellets using a CEP-4 probe (Qbiogene, Illkirsch, France) Results: Karyotypes were available for 37cases (2 failures): 24 were normal, 3 had a trisomy 4, (2 as the sole abnormality), 2 a partial loss of 17p (one deletion and one additional material), 2 a loss of sexual chromosome, 1 a t(11;18), 1 a partial duplication of 17q, 1 a partial monosomy 5q with insertion of unknown material in 4q, 1 a complex karyotype with a partial trisomy 4 and 2 with abnormal independent clones. IP-FISH: The 3 complete trisomies 4, already seen by conventional cytogenetic, were confirmed and 4 others were detected. The mean percentage of cells with trisomy 4 was 31% (11–52%) (cut-off 2%, median+4SD) Discussion: We found that 7 of 39 patients (18%) with WM harboured a complete trisomy 4 with IP-FISH, contrary to only three (8%) with conventional cytogenetic. This study, performed in interphase, is independent of the proliferation rate. Therefore the prevalence is probably underestimated since IP-FISH was realized in unselected cells: in 11 patients the percentage of clonal involvement of the bone marrow was under 30% and it was unknown in 6 patients. The mean percentage of bone marrow lymphoplasmocytic cells was higher for patients with trisomy 4 (63%) than in others (43%). The trisomy 4 was the only abnormality in 2 of the 3 karyotypes where it was identified and in one sporadic case reported previously. Thus it could be a specific primary genetic event in the WM. Moreover, the detection of one partial trisomy 4 define an interesting region with potential oncogene. cKIT, located at 4q12, was a good candidat gene and molecular studies are in process. Conclusion: We identified a new recurrent chromosomal abnormality in WM, the trisomy 4, with a prevalence of at least 18%. The exact prevalence of this abnormality should be determined by IP-FISH study in CD19+ and CD138+ selected cells. This abnormality could be primary and could lead to elusive the molecular pathogenesis of WM.