Leukemia (1999) 13, 460–468 1999 Stockton Press All rights reserved 0887-6924/99 $12.00 http://www.stockton-press.co.uk/leu Translocations involving the short arm of chromosome 17 in chronic B-lymphoid disorders: frequent occurrence of dicentric rearrangements and possible association with adverse outcome E Callet-Bauchu1, G Salles2, S Gazzo1, C Poncet1, D Morel1, J Page`s1, B Coiffier2, P Coeur1 and P Felman1 1Laboratoire Central d’He´matologie and 2Service d’He´matologie Clinique, Centre Hospitalier Lyon Sud, France, and UPRES-JE 1879 ‘He´mopathies Lymphoı¨des Malignes’, Universite´ Claude Bernard, Pierre-Be´nite, France Unbalanced translocations involving chromosome arm 17p, came from a FISH study demonstrating TP53 deletions in 11 where the TP53 tumor suppressor gene localizes, are rarely out of 90 (12%) patients with B-CLL.10 More recently, using described in chronic lymphocytic leukemia and small lympho- cytic lymphoma (CLL/SLL), but recent use of molecular cyto- FISH analysis, we studied three CLL/SLL cases with a recurrent genetic techniques have indicated a significant incidence of t(4;17) translocation and found that the rearrangement had TP53 deletions, suggesting the involvement of chromosome retained the centromeres of both chromosomes, leading to the 17p in these disorders. By conventional karyotype, we have formation of a dicentric rearrangement and to the TP53 identified unbalanced translocations involving 17p in 14 out of monoallelic deletion.11 Those structural abnormalities of chro- 123 (11%) CLL/SLL patients with clonal abnormalities. Cases mosome 17 with loss of material from the short arm through were characterized by resistance to chemotherapy and a poor clinical outcome. The karyotypes presented a high incidence dicentric translocations have been rarely described in chronic of complex rearrangements and 17p translocations were lymphoproliferative disorders. characterized by various partners. In 10 cases a centric fusion In the present study, we focused our interest on 17p translo- was assessed by fluorescent in situ hybridization (FISH) experi- cations in patients with CLL/SLL. We identified 11 additional ments using specific centromeric probes. The incidence of cases with rearrangements involving chromosome 17p and dicentric translocations in these series is therefore signifi- various other partners. To further characterize the chromo- cantly higher than usually described, arising in up to 71% (10 out of 14 cases). In all cases, translocations led to a monosomy somal abnormalities, we investigated the cases by means of 17p and to a TP53 monoallelic deletion. The adverse clinical FISH using painting probes, centromeric probes and a specific outcome confirms that structural abnormalities involving chro- TP53 DNA probe. We also analyzed the outcome of patients mosome 17p are associated with disease progression in with tumoral cells carrying the 17p abnormalities. patients with chronic lymphoproliferative disorders. Keywords: chromosome 17; dicentric translocation; FISH; chronic B cell disorders Materials and methods Introduction Patients In B cell chronic lymphocytic leukemia and small lympho- From February 1993 to October 1997, 226 patients with cytic lymphoma (CLL/SLL), clonal chromosome abnormalities CLL/SLL were referred to our institution. Among them, 123 are identified in approximately 50% of patients.1 However, presented clonal chromosome abnormalities. In 14 cases, kar- conventional karyotype analysis has been often hampered by yotypes displayed translocations involving the short arm of the low in vitro mitotic activity of the tumoral cells.2 Recent chromosome 17. Clinical and biological data of three patients improvement in fluorescence in situ hybridization (FISH) tech- have been previously reported.11 All cases were reviewed by niques has provided an alternative approach to assess the cyto-histopathologist experts. In 11 patients (eight males and detection and the true incidence of chromosomal aberrations three females) the diagnosis of B-CLL was assessed by the in those hematological malignancies.3,4 Among them, trisomy presence of a persistent lymphocytosis greater than 5 × 109/l. 12 and structural abnormalities of chromosome bands 13q14, In three cases, the diagnosis of SLL was based on clinical and 14q32 and 11q22–23 are the most common aberrations cyto-histological findings: patients presented at diagnosis with reported and may be associated with adverse clinical fea- disseminated lymph node enlargement, hepatomegaly and tures.5–7 In addition, in SLL (including leukemic forms), a splenomegaly while the peripheral white blood counts were del(6)(q21q23) has also been documented as a frequent recur- normal. In all cases, the tumoral cells displayed co-expression rent chromosome abnormality.8 of the CD5, CD19 and CD23 antigens, an immunophenotype In most conventional banding analysis of CLL/SLL, aber- in agreement with the diagnosis of CLL/SLL. All medical rec- rations involving chromosome 17 have not been identified as ords were reviewed for treatments and outcome, and response frequent abnormalities. In the Second International Working was assessed using the revised NCI criteria.12 Party on Chromosomes in CLL (IWCCLL), which compiled the cytogenetic data from 662 patients, 22 of 649 (4%) evaluable tumors had rearrangements of chromosome 17, mostly trans- locations of 17p, where the TP53 tumor suppressor gene Chromosome studies localizes.9 Further information about involvement of 17p Conventional cytogenetic studies were performed on periph- eral blood, bone marrow or lymph nodes samples. Chromo- Correspondence: E Callet-Bauchu, Unite´ d’He´matologie Cellulaire, Laboratoire Central d’He´matologie, Centre Hospitalier Lyon Sud, some analyses were carried out on RHG-banded metaphases 69495 Pierre-Be´nite Cedex, France; Fax: 33 04 78 86 33 40 as previously described and evaluated according to the ISCN Received 29 May 1998; accepted 23 October 1998 (1995) recommendations.13 Chromosome 17 dicentric rearrangements in chronic B cell disorders E Callet-Bauchu et al 461 Metaphase and interphase FISH abnormal metaphases were in a tetraploid range (Table 1). Complex karyotypes with multiple abnormal clones were Dual-color chromosome painting: For FISH study, air- observed in 11 of the 14 patients. Cells with a normal karyo- dried slides with conventional chromosome preparations were type were identified in all cases except three (patients 2, 7 used. Hybridization experiments were performed as pre- and 12). Partial karyotypes illustrating structural rearrange- viously reported with minor modifications.11 The probes used ments involving chromosome 17 are shown in Figures 1 were a biotin-labeled chromosome 17 probe (Oncor, Gai- and 2. thersburg, MD, USA) and various digoxigenin-labeled paints By conventional cytogenetic analysis, seven patients specific for chromosomes 2, 4, 12, 13, 15, 18, 19, 20 and presented a derivative chromosome 17 resulting from a trans- 22 (Oncor). The probes cohybridized to metaphases at 37°C location involving 17p11 and 2q11 (patient 1), 13q11 (patient overnight, and slides were washed in 0.5 × SSC at 72°C for 2), 15q15 (patient 3), 18q11 (patients 4–6), and either 15q11, 5 min, followed by 1 × phosphate-buffered detergent (PBD; 20q11 or 22q11 in patient 7 for whom a jumping translo- Oncor) for 2 min at room temperature. Hybridization signals cation was identified. Moreover, in patient 3 the derivative were detected by the use of a layer of fluorescein isothiocyan- chromosome was observed in two related clones, leading to ate (FITC)-conjugated avidin antibodies (5 ␮g/ml; Vector Lab- a der(17)t(15;17) in the stemline clone and to a oratories, Burlingame, CA, USA) in combination with rhoda- der(17)t(4;17)t(15;17) in an additional deviating subclone. mine-labeled antidigoxigenin antibodies (2 ␮g/ml; Boehringer In the remaining seven patients the consistent cytogenetic Mannheim, Germany). Slides were then counterstained with abnormality resulted in loss of a chromosome 17 and the pres- DAPI (0.1 ␮g/ml) in an antifade solution, observed and photo- ence of unbalanced rearrangements involving chromosome graphed on a Zeiss epifluorescence axioskop equipped with 17, in a segment distal to 17q11 and either chromosome 12p the appropriate filter combinations using a Kodak ektachrome (patients 8 and 9), 12q (patient 10), 19q (patient 11) or 4p 400 film. (patients 12–14). In patients 8 and 9, the derivative chromo- In a few cases, a second stage hybridization using somes 12 were interpreted as der(12)t(12;17)(p12;q11). In additional paints was required. This prompted us to investi- patient 10, the derivative chromosome 12 was generated by gate the same slide repeatedly by applying various combi- two translocations, one involving the short arm, resulting from nations of biotin- or digoxigenin-labeled probes. Consecutive a translocation between band 12p13 and the segment experiments were then performed as described above. Signals 18q24 → 18 qter and the other involving the long arm, with related to probes applied in previous hybridizations were not a translocation of the segment 17q11→17 qter to band 12q15. visible using adequate labeled probes and detection systems In patient 11, the same chromosome 17 was involved in two or fluorochromes, as recently reported.14 distinct clones leading to a der(19) t(17;19)(q11;q13) in the stemline clone and to a der(2) t(2;17)(p24;q11) in an additional unrelated clone. FISH with ␣ satellite DNA probes: Dual-color hybridiz- Associated clonal abnormalities were identified in