Conventional cytogenetic analysis in B-cell chronic lymphocytic leukemia (B-CLL) has been very difficult, and the prognostic significance of specific chromosome aberrations is under discussion. Recent improvements in fluorescence in situ hybridization (ISH) techniques have provided an alternative approach for the detection of chromosome aberrations. Here, an interphase cytogenetic study was performed to analyze the incidence and prognostic significance of a p53 gene deletion in B-CLL and related disorders. We studied mononuclear cells from 100 patients with chronic B-cell leukemias [B-CLL, 90 patients; B-prolymphocytic leukemia (B-PLL), 7; Waldenstrom's macroglobulinemia (WM), 3] by fluorescence ISH with a genomic p53 DNA probe. In a subset of patients, additional G-banding analysis and single strand conformation polymorphism (SSCP) analysis was performed. Seventeen of the 100 patients [17%; B-CLL, 11 of 90 (12%); WM, 1 of 3; B-PLL, 5 of 7] exhibited a monoallelic p53 gene deletion by ISH. G- banding analysis demonstrated abnormalities of chromosome 17 in 13 of these 17 patients, all leading to loss of band 17p13. SSCP analysis showed aberrant bands in 9 of 14 patients with a p53 gene deletion. None of 12 patients with a p53 gene deletion compared with 20 of 36 patients (56%) without a deletion responded to therapy with fludarabine or pentostatin (P < .001). The difference in survival probabilities from the time of diagnosis and from the start of treatment with purine analogs between the two groups was highly significant (P < .001). In multivariate analysis, p53 gene deletion was the strongest prognostic factor for survival. In conclusion, p53 gene deletion predicts for non- response to therapy with purine analogs and for poor survival in chronic B-cell leukemias.

The presence of BCR-ABL fusion genes has important diagnostic and prognostic implications in chronic myeloid leukemia (CML) and acute lymphoblastic leukemia (ALL). The CML-specific chimeric BCR-ABL gene with a break involving the major breakpoint cluster region (M-bcr) of the BCR-gene has been detected by means of fluorescence in situ hybridization (FISH). In this study, we present a FISH protocol that allows the detection of breaks in both the major and the minor breakpoint cluster region (m-bcr). Three hybridization signals of D107F9, a yeast artificial chromosome (YAC)-derived probe spanning the breakpoint regions of the BCR gene, were indicative of the translocation events. To increase the specificity further, this probe was combined with cos-abl 8, a cosmid probe flanking the breakpoint within the ABL gene for dual-color hybridization. Samples of 21 patients with CML, the ALL-derived cell line SUP-B15, and of seven patients with Philadelphia chromosome (Ph1)-positive ALL (three of them with breakpoints within m-bcr) were examined. BCR-ABL fusion was detected in all cases with high specificity (false-positive nuclei: mean, 0.1%). On cytogenetic preparations, the percentages of BCR-ABL- positive interphase cells ranged from 53% to 91%. Comparable efficiencies were achieved on blood smears. In conclusion, hybridization with D107F9 and cos-abl 8 allows unambiguous diagnosis of BCR-ABL genes and is likely to become an important tool for the monitoring of therapies in patients with CML and ALL.

Deletions or translocations of 13q, most commonly involving band 13q14, belong to the most frequent structural chromosome abnormalities in B-cell chronic lymphocytic leukemia (B-CLL). In a combined metaphase and interphase cytogenetic study using conventional G-banding analysis and fluorescence in situ hybridization (ISH) we previously analysed the retinoblastoma susceptibility gene (RB-1) and its chromosomal locus 13q14 in 35 patients with chronic B-cell leukemias. We report here on the interphase cytogenetic analysis of 109 cases with chronic B-cell leukemias [B-CLL = 90; B-prolymphocytic leukemia (B-PLL) = 6, hairy cell leukemia (HCL) = 13]; a subset of 49 patients (B-CLL = 45; B-PLL = 4) was studied by conventional G-banding analysis. By G-banding, 5/45 (11%) patients with B-CLL had deletions or translocations affecting band 13q14; in contrast, ISH to interphase cells showed RB-1 deletion in 19/90 (21%) patients with B-CLL. No 13q14 abnormalities or RB-1 deletion were detected in patients with B-PLL and HCL. Our data confirm the high frequency of RB-1 deletions in B-CLL and further emphasize the possible pathogenetic role of this genomic region.

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