Genomic Aberrations In Bcll

Two major subjects can be differentiated with respect to the genetic analysis of B-CLL: on the one hand, genomic aberrations, which, as acquired changes, may be involved in the initiation and progression of the disease, and, on the other hand, the mutation status of the variable segments of immunoglobulin heavy chain genes (VH), which may reflect the cellular origin of B-CLL.

Since the early 1980s, chromosome banding analyses of malignant B-cells have been performed using B-cell mitogens (11-18). Up to the early 1990s, clonal aberrations could be dem-

From: Contemporary Hematology Chronic Lymphocytic Leukemia: Molecular Genetics, Biology, Diagnosis, and Management Edited by: G. B. Faguet © Humana Press Inc., Totowa, NJ 57

Fig. 1. Marked cervical and axillary lymphadenopathy in a B-CLL patient with deletion 11q. (From ref. 9, with permission.)

MONTHS

Fig. 2. Estimated survival times of B-CLL subgroups according to Rai stage (from ref. 6, with permission).

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Fig. 2. Estimated survival times of B-CLL subgroups according to Rai stage (from ref. 6, with permission).

onstrated in only 40-50% of B-CLL cases using chromosome banding (19-21). Frequently, and despite the use of B-cell mitogens, nonclonal T-cells without chromosomal aberration were analyzed (22). More recently, the development of molecular cytogenetic techniques like fluorescence in situ hybridization (FISH) has led to considerable improvement in the diagnostics of genetic aberrations in tumor cells (23,24). FISH allows sensitive detection of specific sequences in the genome using cloned DNA fragments as probes. Signal number and location reflect numerical and structural changes of the corresponding chromosomal regions. The ability to detect aberrations not only on metaphase chromosomes but also in interphase cell nuclei is of great importance, especially in B-CLL (interphase cytogenetics; Fig. 3) (25). Interphase cytogenetic studies using FISH showed that the incidence of genomic aberrations in B-CLL was markedly

Fig. 3. Interphase FISH in B-CLL. (A) 11q deletion as demonstrated by the single red signal in five of the six nuclei shown. Two green signals of an internal control probe prove a high hybridization efficiency. The single cell with two red signals probably represents a nonleukemic cell from the specimen. (B) Biallelic deletion at 13q. Two of the three nuclei show no red hybridization signal of a probe containing marker D13S272, demonstrating biallelic loss of this region, whereas an adjacent probe containing marker D13S273 is retained in a disomic fashion. The single cell with two red and two green signals probably represents a nonleukemic cell. (C) Trisomy 12q (three green hybridization signals) and monoallelic deletion 13q14 (single red signal) in two of three nuclei in a B-CLL specimen. A single cell reflecting the normal disomic status of the two regions is shown for comparison. (From ref. 27, with permission.)

Fig. 3. Interphase FISH in B-CLL. (A) 11q deletion as demonstrated by the single red signal in five of the six nuclei shown. Two green signals of an internal control probe prove a high hybridization efficiency. The single cell with two red signals probably represents a nonleukemic cell from the specimen. (B) Biallelic deletion at 13q. Two of the three nuclei show no red hybridization signal of a probe containing marker D13S272, demonstrating biallelic loss of this region, whereas an adjacent probe containing marker D13S273 is retained in a disomic fashion. The single cell with two red and two green signals probably represents a nonleukemic cell. (C) Trisomy 12q (three green hybridization signals) and monoallelic deletion 13q14 (single red signal) in two of three nuclei in a B-CLL specimen. A single cell reflecting the normal disomic status of the two regions is shown for comparison. (From ref. 27, with permission.)

underestimated in banding studies (9). In B-CLL cases with abnormal karyotype by banding analysis but clonal aberrations by interphase FISH, the metaphase cells are derived from nonclonal T-cells and therefore do not reflect the karyotype of the malignant B-CLL cells.

Among the genomic aberrations whose incidence has been underestimated in B-CLL in banding studies are particularly the deletions of bands 13q14 and 11q22-q23, (9); while trisomy 12, which was originally described as the most frequent aberration of B-CLL in studies using chromosome banding, has been rated the third most frequent aberration by interphase FISH (9,26,27). In a study on 325 B-CLL patients, a comprehensive disease-specific probe set was used to detect the most important genomic gains, like partial trisomies 12q13, 3q27, and 8q24 and the most frequent genomic losses in bands 13q14, 11q22-q23, 6q21, 6q27, and 17p13 and translocations in band 14q32 using FISH (9) (Table 1). Overall genomic aberrations were found in more than 80% of all cases. The most frequent aberration by far was deletion of band 13q14, which was found in 55% of the cases. Other frequent aberrations were deletion 11q22-q23 (18%), trisomy 12q13 (16%), deletion 17p13 (7%), deletion 6q21 (6%), trisomy 8q24 (5%), translocation 14q32 (4%), and trisomy 3q27 (3%). Somewhat more than half of the cases showed only a single aberration; one-fifth of the cases showed two and nearly one-tenth showed more than two aberrations.

This precise determination of the incidence of chromosomal aberrations provides the basis for further studies of the role of these changes in the pathogenesis and progression of the disease. Thus, genes assumed to be involved in the pathogenesis of B-CLL could be identified by physical mapping of the minimal affected regions and by the strategy of positional cloning as well as the analysis of candidate genes (for review, see ref. 27).

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