At diagnosis, approximately half of myelodysplastic (MDS) patients presents a normal karyotype by conventional cytogenetic analysis (CCA). Fluorescent in situ hybridization (FISH) is more sensitive than CCA allowing for the detection of minor clones and of submicroscopic lesions. We have analyzed by FISH 101 MDS patients with normal karyotype for the occurrence of the abnormalities which are most frequently observed in MDS (ie ؊5/5q؊, ؊7/7q؊, ؉8, 17p؊). In 18 patients, 15 to 32% of interphase cells were found to carry one FISH abnormality. Six patients presented trisomy 8, five had del(5)(q31), five del(7)(q31), one monosomy 7 and one del(17)(p13). FISH abnormalities were more frequently observed among patients with an increased percentage of bone marrow blasts (P ‫؍‬ 0.001). FISH abnormalities were also associated with a higher rate of progression into AML (13/18 vs 12/83, P Ͻ 0.001) and were predictive for a worse prognosis (P Ͻ 0.001). Multivariate analysis indicated that FISH positivity and IPSS risk group were independent predictors for a poor survival (P ‫؍‬ 0.0057 and 0.0123, respectively) and for leukemic transformation (P ‫؍‬

Summary.To better define the role of exposure to myelotoxic agents in the genesis of myelodysplastic syndrome (MDS), we carried out (a) a case-control study for the determination of the relative risk (RR) of developing MDS, including 178 consecutive patients and 178 sex-and age-matched controls; (b) a study of clinicobiological features in MDS arising after occupational exposure to myelotoxic agents and in MDS in 'non-exposed' patients. The definition of the 'exposure' status was based on a predetermined questionnaire, with calculation of an 'exposure' index (hours/ day × days/year × years). Cumulative exposure to pesticides or to organic solvents, for >2400 h, was recorded in 48 and 25 MDS patients, respectively, compared to 27 and four controls (P < 0·00001; RR 3·74; 95% confidence interval 2·02-5·37). Older age and an excess of refractory anaemia with ringed sideroblasts and refractory anaemia with excess of blasts was noted among 'exposed' MDS-patients (group 1), compared to non-exposed MDS-patients (group 2). 68·3% patients in group 1 had clonal chromosome changes, compared with 43·2% patients in group 2. Complex karyotypes, ¹7/7q¹, ¹5/5q¹, þ8, 7p and 17p aberrations were seen more frequently in group 1, whereas a normal karyotype, isolated 5q¹ or 20q¹ occurred more frequently in group 2. The association of exposure to myelotoxic agents with older age at presentation and with unfavourable chromosome changes accounted for the shorter survival observed in 'exposed' patients. These data show that occupational exposure to pesticides and organic solvents in our region resulted in an increased RR of developing MDS and that a distinct cytogenetic profile was associated with MDS in 'exposed' patients. These findings provide strong indirect evidence that these agents may play a role in the pathogenesis of MDS, preferentially targeting some of the chromosome regions which are frequently involved in therapy-related myeloid neoplasias.

In order to define better the cytological and clinical features of atypical B-cell chronic lymphocytic leukaemia (B-CLL) with t(11:14)(q13;q32), sequential morphologic immunological and cytogenetic studies were performed in seven patients belonging to a series of 72 consecutive cases presenting with a diagnosis of CLL or atypical CLL according to the FAB criteria. Cytologic diagnosis in these seven patients with t(11;14) was typical CLL in two cases presenting with < 10% large lymphocytes (LL) and prolymphocytes (PL) and atypical CLL in five cases in which LL and PL comprised between 10% and 55%. The diagnosis was supported by histologic findings on bone marrow biopsy (five cases) or splenectomy specimens (two cases). A progressive increase of peripheral LL and PL was observed, resulting in a switch of FAB diagnosis over a 6-60-month period from typical CLL into atypical CLL in two cases and from atypical CLL into prolymphocytic leukaemia in five cases. Immunophenotyping showed a mature B-cell phenotype with CD19, CD22, CD24 positivity and CD10 negativity in all patients. A bright-staining pattern for surface immunoglobulins (SIg) was detected in 6/7 cases, CD5 positivity in 6/7 cases, and CD23 positivity in 1/7 cases. The FMC-7 monoclonal antibody was positive in > 40% cells in 5/6 cases. Chromosome changes in addition to t(11;14) were seen in five cases; in two cases unbalanced translocations involving the 3q21 chromosome region, resulting in partial trisomy for the long arm of chromosome 3, were detected early in the course of the disease. Karyotype evolution that was associated with disease progression occurred in 3/6 assessable patients. Comparison of these findings with similar data from 65 B-CLL patients without t(11:14) showed that atypical morphology, switch of FAB diagnosis during the course of the disease, and karyotype evolution were more frequently seen in cases with t(11;14) (5/7 v 15/65 cases, P = 0.015, 7/7 v 7/65 cases, P < 0.0001, and 3/6 v 5/45 assessable cases, P = 0.04, respectively). The frequency of positivity for CD23 and bright SIg staining differed significantly in the two groups. It is concluded that t(11;14) identifies a cytologically atypical subset of B-CLL, characterized by frequent cytologic and cytogenetic evolution and by a distinct immunological profile, sharing some biological features with mantle cell lymphoma.

Clinicobiological, histological, cytogenetic and molecular genetic studies were performed in a case of atypical B-cell chronic lymphocytic leukaemia (B-CLL) with the t(11;14)(q13;q32) evolving into Richter's syndrome (RS) in order (a) to determine the clonal relationship between the cell of origin for B-CLL and RS, and (b) to analyse genetic events underlying the disease progression in this patient. After 4 years following diagnosis, a rapid deterioration of the clinical picture occurred, concomitant with the appearance of large lymphoid blasts in peripheral blood (PB), bone marrow (BM) and ascites samples. A diagnosis of RS was made and cytogenetic analysis revealed karyotype evolution with trisomy 7 and del(17p) in addition to t(11;14). Fluorescence in situ hybridization showed 78% lymphoid blast cells obtained from ascites sample to be trisomic using a chromosome-7-specific pericentromeric probe. Whereas no rearrangement of the c-myc proto-oncogene was detected at disease progression, direct sequencing of p53 gene exon 5-9 revealed an exon 7 missense point mutation. This abnormality was not present in the CLL phase. Immunological staining with the monoclonal antibody PAb-1801, detecting the p53 protein product, revealed a negative pattern in the CLL phase, whereas 24% positivity was documented in representative samples obtained at RS. It is concluded that RS was cytogenetically related with B-CLL in this patient, suggesting the occurrence of a bona fide transformation and that the mutation of p53 exon 7, in association with the development of 17p deletion, possibly played a role in the development of RS.

To define better the chromosomal profile of atypical chronic karyotype may in fact carry chromosomally abnormal clones lymphocytic leukemia (aCLL), cytogenetic and interphase cytoescaping detection at metaphase analysis due to their low genetic studies were performed in 43 cases, using mitogenmitotic index. a relatively poor prognosis may be associated with aCLL.The presence of ؉12, 13q14 deletions, 11q, and 6q21-q23 anomalies in 19 cases was associated with a 2-month median interTo define better the cytogenetic profile of aCLL, this study val between diagnosis and start of treatment, as compared with of 43 cases was carried out combining conventional chromoa 24-month median interval in 14 cases with normal karyotype some analysis (CCA) and FISH, using probes detecting the two or non-recurrent chromosome changes (P ‫؍‬ 0.003). We conmost frequent anomalies in CLL, ie +12 and del(13q14).clude that aCLL is characterized by a relatively high incidence of chromosome anomalies, with recurrent chromosome changes, involving chromosomes 12, 13q14, 6q21q23, 11q, selected on the basis of the presence of more than 10% large Keywords: atypical CLL; cytogenetics; FISH lymphocytes and/or prolymphocytes (see below) among approximately 260 B-CLLs, seen at our Institution over a 9-year period. Materials and methods IntroductionDiagnoses were made in all cases according to standard clinical, cytologic and immunologic criteria. Histologic studThe importance of cytogenetics of B cell chronic lymphocytic ies were performed for diagnostic purposes in 27 cases (bone leukemia (CLL) has been highlighted over the past 10 years biopsy in 19 cases and superficial lymph node biopsy in eight in a number of studies, reviewed by Juliusson and Gahrton, 1 cases). Transformation of aCLL into prolymphocytic leukemia documenting that some chromosome changes, such as tri-(PLL) was not an exclusion criteria, whereas those patients somy 12, del (11q) or 14q anomalies are associated with therpresenting with 'de novo' PLL were not considered in this apy-demanding disease and, ultimately, with shorter survival.