Interstitial 13q14 deletions detected in the karyotype and translocations with concomitant deletion at 13q14 in chronic lymphocytic leukemia: Different genetic mechanisms but equivalent poorer clinical outcome

Deletion of 13q14 as the sole abnormality is a good prognostic marker in chronic lymphocytic leukemia (CLL). Nonetheless, the prognostic value of reciprocal 13q14 translocations [t(13q)] with related 13q losses has not been fully elucidated. We described clinical and biological characteristics of 25 CLL patients with t(13q), and compared with 62 patients carrying interstitial del(13q) by conventional G‐banding cytogenetics (CGC) [i‐del(13q)] and 295 patients with del(13q) only detected by fluorescence in situ hybridization (FISH) [F‐del(13q)]. Besides from the CLL FISH panel (D13S319, CEP12, ATM, TP53), we studied RB1 deletions in all t(13q) cases and a representative group of i‐del(13q) and F‐del(13q). We analyzed NOTCH1, SF3B1, and MYD88 mutations in t(13q) cases by Sanger sequencing. In all, 25 distinct t(13q) were described. All these cases showed D13S319 deletion while 32% also lost RB1. The median percentage of 13q‐deleted nuclei did not differ from i‐del(13q) patients (73% vs. 64%), but both were significantly higher than F‐del(13q) (52%, P < 0.001). Moreover, t(13q) patients showed an increased incidence of biallelic del(13q) (52% vs. 11.3% and 14.9%, P < 0.001) and higher rates of concomitant 17p deletion (37.5% vs. 8.6% and 7.2%, P < 0.001). RB1 involvement was significantly higher in the i‐del(13q) group (79%, P < 0.001). Two t(13q) patients (11.8%) carried NOTCH1 mutations. Time to first treatment in t(13q) and i‐del(13q) was shorter than F‐del(13q) (67, 44, and 137 months, P = 0.029), and preserved significance in the multivariate analysis. In conclusion, t(13q) and del(13q) patients detected by CGC constitute a subgroup within the 13q‐deleted CLL patients associated with a worse clinical outcome. © 2014 Wiley Periodicals, Inc.     

Influence of clone and deletion size on outcome in chronic lymphocytic leukemia patients with an isolated deletion 13q in a population‐based analysis in British Columbia,Canada

Deletion of the long arm of chromosome 13 (del(13q)) as the sole abnormality in chronic lymphocytic leukemia (CLL) portends a good prognosis; however, there is great outcome heterogeneity within this subgroup. The percentage of cells with a del(13q) (clone size) and the extent of the deletion are two factors that may affect outcome in CLL patients with isolated del(13q). We analyzed 248 CLL patients from the BC Provincial CLL database identified as having isolated del(13q) detected pretreatment by interphase fluorescence in situ hybridization to determine what impact clone and deletion size had on overall survival (OS) and treatment free survival (TFS). Patients with 60% or more of nuclei with a del(13q) had shorter TFS and shorter OS. A large deletion, encompassing the RB1 gene locus, was detected in half of the 90 cases with available specimens for testing, and there was no significant difference in OS and TFS between RB1‐deleted and RB1‐not‐deleted cases. Further study in a larger sample size is required to determine the clinical interest of RB1 locus testing; however, clone size of del(13q) does predict TFS and OS and may better refine prognosis in this clinically heterogeneous population. © 2015 Wiley Periodicals, Inc.     

Prognostic relevance of monosomy at the 13q14 locus detected by fluorescence in situ hybridization in B-cell chronic lymphocytic leukemia

Deletion of the chromosome band 13q14, which contains the putative deleted in B-cell malignancy (DBM) gene, and trisomy 12 have been demonstrated by fluorescence in situ hybridization (FISH) techniques in malignant B-cells from patients with B cell chronic lymphocytic leukemia (B-CLL). However, the prognostic relevance of 13q14 abnormalities as detected by FISH is unknown. We prospectively studied malignant blood cells from 54 consecutive, untreated B-CLL patients using FISH probes to the RB1 locus and DBM (markers D13S25 and D13S319) for band 13q14, as well as probes to chromosome 12. The cells from all cases were CD5+ CD20+, expressed clonally restricted surface immunoglobulin light chain, and had typical features for B-CLL on careful blood smear morphologic evaluation. Patients were followed for a mean of 3.9 years and treatment-free survival (TFS) was used in the prognostic factor analysis. Twenty-four (44%) patients were observed to have monosomy of the RB1 locus and 26 (48%) monosomy of D13S25 and D13S319. The 26 patients who had a deletion at at least one of these loci had a 48.4 month (mo) median TFS vs 31.1 mo for those without evidence of deletion at any 13q14 locus (p = 0.07). The seven patients found to have trisomy 12 had a median TFS of 6.9 mo vs 39.3 mo for those diploid for chromosome 12 (p < 0.01). When these seven patients with trisomy 12 were excluded from the analysis, patients who had a deletion at 13q14 tended to have a longer median TFS (50.1 vs 36.2 mos), but this was not statistically significant (p = 0.2). This study confirms the prevalence of 13q14 deletions in B-CLL and suggests that patients with this abnormality have a better TFS than those with trisomy 12.     

应用荧光原位杂交技术检测84例慢性淋巴细胞白血病遗传学异常及预后分析

 摘要 目的：探讨荧光原位杂交 (fluorescence in situ hybridization,FISH) 技术检测慢性淋巴细胞白血病(chronic lymphocytic leukemia,CLL)患者的遗传学异常,并分析与其他预后指标的相关性及预后价值。方法：回顾性分析采用FISH技术进行着丝粒探针CSP12(12p11.1～12q11.1)和序列特异性探针D13s25(13q14.3)、ATM (11q22.3)、RB1(13q14)、p53(17p13.1)检测过的84例CLL患者的临床以及实验室资料。分析FISH技术检测CLL患者中遗传学异常的发生率,并分析遗传学异常与CLL患者年龄、性别、Binet分期、血清乳酸脱氢酶(lactate dehydrogenase,LDH)、CD38表达、生存期之间的相关性。结果：(1)84例CLL患者中,62例有至少有一种遗传学异常。遗传学异常的检出率为73.8%。(2)共发现5种遗传学异常,依次为13q14缺失（56%）、P53缺失（28.6%）、-12（16.7%）、ATM缺失（13.1%）、+12（13.1%）。其中-12为CLL病人中新发现遗传学异常。(3)分析上述5种遗传学异常与年龄、性别、Binet分期、LDH水平、CD38表达的关系,在>60岁年龄病人里,+12阳性率显著高于<60岁病人(24%VS3%, P=0.042),与性别、Bient分期、LDH水平、CD38表达无关。(4)在LDH增高组中,CD38高表达的病例比例增高（67%VS20%, P=0.015）。(5) 伴有del(17p13)或del(11q22)或复杂异常中位生存时间为75个月,单纯具有del(13q14)异常组的中位生存时间为150个月,其他中位生存时间为120月,但两两比较没有达到统计学差异。结论：(1)FISH技术是一种在分析CLL患者遗传学异常方面较为快速、准确和敏感的方法; (2)CLL病人中除了常见13q14缺失、P53缺失、ATM缺失、+12之外,还可以出现-12异常。     

Dual-probe fluorescence in situ hybridization assay for detecting deletions associated with VCFS/DiGeorge syndrome I and DiGeorge syndrome II loci

Over 90% of patients with DiGeorge syndrome (DGS) or velocardiofacial syndrome (VCFS) have a microdeletion at 22q11.2. Given that these deletions are difficult to visualize at the light microscopic level, fluorescence in situ hybridization (FISH) has been instrumental in the diagnosis of this disorder. Deletions on the short arm of chromosome 10 are also associated with a DGS-like phenotype. Since deletions at 22q11.2 and at 10p13p14 result in similar findings, we have developed a dual-probe FISH assay for screening samples referred for DGS or VCFS in the clinical laboratory. This assay includes two test probes for the loci, DGSI at 22q11.2 and DGSII at 10p13p14, and centromeric probes for chromosomes 10 and 22. Of 412 patients tested, 54 were found to be deleted for the DGSI locus on chromosome 22 (13%), and a single patient was found deleted for the DGSII locus on chromosome 10 (0. 24%). The patient with the 10p deletion had facial features consistent with VCFS, plus sensorineural hearing loss, and renal anomalies. Cytogenetic analysis showed a large deletion of 10p [46, XX,del(10)(p12.2p14)] and FISH using a 10p telomere region-specific probe confirmed the interstitial nature of the deletion. Analysis for the DGSI and the DGSII loci suggests that the deletion of the DGSII locus on chromosome 10 may be 50 times less frequent than the deletion of DGSI on chromosome 22. The incidence of deletions at 22q11.2 has been estimated to be 1 in 4000 newborns; therefore, the deletion at 10p13p14 may be estimated to occur in 1 in 200,000 live births.     

Dual‐probe fluorescence in situ hybridization assay for detecting deletions associated with VCFS/DiGeorge syndrome I and DiGeorge syndrome II loci

Over 90% of patients with DiGeorge syndrome (DGS) or velocardiofacial syndrome (VCFS) have a microdeletion at 22q11.2. Given that these deletions are difficult to visualize at the light microscopic level, fluorescence in situ hybridization (FISH) has been instrumental in the diagnosis of this disorder. Deletions on the short arm of chromosome 10 are also associated with a DGS‐like phenotype. Since deletions at 22q11.2 and at 10p13p14 result in similar findings, we have developed a dual‐probe FISH assay for screening samples referred for DGS or VCFS in the clinical laboratory. This assay includes two test probes for the loci, DGSI at 22q11.2 and DGSII at 10p13p14, and centromeric probes for chromosomes 10 and 22. Of 412 patients tested, 54 were found to be deleted for the DGSI locus on chromosome 22 (13%), and a single patient was found deleted for the DGSII locus on chromosome 10 (0.24%). The patient with the 10p deletion had facial features consistent with VCFS, plus sensorineural hearing loss, and renal anomalies. Cytogenetic analysis showed a large deletion of 10p [46,XX,del(10)(p12.2p14)] and FISH using a 10p telomere region‐specific probe confirmed the interstitial nature of the deletion. Analysis for the DGSI and the DGSII loci suggests that the deletion of the DGSII locus on chromosome 10 may be 50 times less frequent than the deletion of DGSI on chromosome 22. The incidence of deletions at 22q11.2 has been estimated to be 1 in 4000 newborns; therefore, the deletion at 10p13p14 may be estimated to occur in 1 in 200,000 live births. Am. J. Med. Genet. 91:313–317, 2000. © 2000 Wiley‐Liss, Inc.     

A der(13)t(7;13)(p13;q14) with monoallelic loss of RB1 and D13S319 in myelodysplastic syndrome

Deletions or translocations of chromosome band 13q14, the locus of the retinoblastoma gene (RB1), have been observed in a variety of hematological malignancies including myelodysplastic syndrome (MDS). We describe here a novel unbalanced translocation der(13)t(7;13)(p13;q14) involving 13q14 in a patient with MDS. A 66-year-old woman was diagnosed as having MDS, refractory anemia with excess of blasts (RAEB-1) because of 7.4% blasts and trilineage dysplasia in the bone marrow cells. G-banding and spectral karyotyping analyses showed complex karyotypes as follows: 46,XX,der(6)t(6;7)(q11;?),der(7)del(7)(?p13)t(6;7)(q?;q11)t(6;13)(q?;q?),der(13)t(7;13)(p13;q14). Fluorescence in situ hybridization (FISH) analyses demonstrated that one allele of the RB1 gene and the microsatellite locus D13S319, located at 13q14 and telomeric to the RB1 gene, was deleted. Considering other reported cases, our results indicate that submicroscopic deletions accompanying 13q14 translocations are recurrent cytogenetic aberrations in MDS. The RB1 gene or another tumor suppressor gene in the vicinity of D13S319, or both, may be involved in the pathogenesis of MDS with 13q14 translocations by monoallelic deletion.     

Delineation of the minimal region of loss at 13q14 in multiple myeloma

Previous studies have focused on the incidence and prognostic implications of 13q14 deletions in multiple myeloma (MM), but none has sought to delineate the minimal common deleted region (CDR). In an effort to do so, dual-color interphase fluorescence in situ hybridization (FISH) was applied on 82 myeloma cases, initially by use of three probes for 13q14 (RB1, D13S319, and D13S25). Deletions were detected in 29/82 (35.4%) cases, and all except one were monoallelic. Subsequently, contiguous YACs, PACs, and a BAC spanning the 13q14-q21 region were employed for deletion mapping in addition to a 13q telomere probe. Large deletions extending to the 13q34 region were found in 55% of the deleted cases, whereas an additional 13.8% showed loss of both 13q34 and 13q14 regions with retention of 13q21. A CDR of approximately 350 kb was identified at 13q14 with the proximal border approximately 120 kb centromeric from D13S319, encompassing an area rich in expressed sequence tagged sites and containing DLEU1, DLEU2, and RFP2 genes. Direct sequencing of the RFP2 gene revealed no mutations in six patients and four MM cell lines harboring deletions of the CDR. However, a role for RFP2 in the pathogenesis of MM cannot yet be excluded, given that alternative mechanisms such as haploinsufficiency remain possible.     

Identification of 13q deletion, trisomy 1q, and IgH rearrangement as the most frequent chromosomal changes found in Korean patients with multiple myeloma

The most frequent genetic aberrations in multiple myeloma (MM) are 13q deletions and translocations involving the immunoglobulin heavy chain gene (IGH). There have been no reports on the cytogenetic abnormalities found in Korean patients with MM. We investigated the actual prevalence and prognostic value of cytogenetic changes using fluorescence in situ hybridization (FISH). FISH studies with 12 different specific probes for the regions containing the genes or chromosome regions (13q, 1q, IGH, p53, MLL, p16, CEP 7, CEP 11, and CEP 12) were performed in 128 patients. The most frequent change found was 13q deletion (48%), followed by trisomy 1q (45%), IGH translocation (37%), and trisomy 11 (26%). Among the three different probes used to detect 13q deletion, D13S25 (48/58) was the most sensitive probe compared to RB (43/58) and D13S319 (39/58). Among the patients showing one or more changes by FISH, 75% (82/110) had a 13q deletion, a trisomy 1q, or an IGH translocation. Azotemia, anemia, thrombocytopenia, intramedullary plasmacytosis, and stage were significantly associated with the 13q deletion; serum beta(2)-microglobulin, thrombocytopenia, and intramedullary plasmacytosis were also related to trisomy 1q. The pattern of molecular cytogenetic changes in Korean patients with MM is somewhat different from what has been observed in reported Caucasian populations: 37 versus 50-70% with regard to the IGH translocation. The prevalence of the 13q deletion was similar in Korean and Caucasian populations, 48 versus 30-50%. We suggest that the detection of at least these three genetic changes, 13q- trisomy 1q, and an IGH rearrangement, would be helpful for follow-up of Korean patients with MM.     

Translocations as a mechanism for homozygous deletion of 13q14 and loss of the ATM gene in a patient with B-cell chronic lymphocytic leukemia

Chromosomal aberrations detected by fluorescence in situ hybridization (FISH) on interphase nuclei are important prognostic markers in chronic lymphocytic leukemia (CLL). Deletions in 13q14 and in 11q22.3 are two of the most frequent aberrations in this disease entity (55 and 18%, respectively) and are usually effected by interstitial deletions. Here, we report on the case of a 66-year-old woman with CLL who was analyzed by conventional metaphase cytogenetics as well as fluorescence in situ hybridization. Deletion-specific probes detected a homozygous loss of two anonymous loci in chromosomal band 13q14 in parallel with a heterozygous loss of the ATM gene located in chromosomal band 11q22.3. Karyotype analysis indicated reciprocal but unbalanced translocations involving chromosomes 3, 11, and 13. Deleted sites on 13q14 appeared to be located within the breakpoint regions of the translocations. We show that mechanisms other than interstitial deletions may lead to loss of critical chromosomal regions in CLL.     

Multiplex ligation-dependent probe amplification and fluorescence in situ hybridization to detect chromosomal abnormalities in chronic lymphocytic leukemia: a comparative study

Chronic lymphocytic leukemia (CLL) is a clinically heterogeneous disease characterized by recurrent chromosomal aberrations of prognostic significance. We aimed to evaluate the potential of the multiplex ligation-dependent probe amplification (MLPA) assay to detect genomic alterations in CLL. Highly purified (>90%) peripheral mononuclear CD19+ cell populations from 100 untreated CLL patients (pts) in early stage disease (Binet stage A) were included in this study. All samples were investigated by fluorescence in situ hybridization (FISH) for the presence of trisomy 12 and 17p13.1, 11q22.3, and 13q14.3 deletions. For MPLA analysis, DNA was amplified by means of two commercially available probes sets allowing the simultaneous screening of 56 genomic sequences. Overall, a high degree of concordance (95%) between MPLA and FISH results was found, if the abnormal clone was present in more than 30% of the leukemic cell population. The use of multiple MPLA probes allowed the fine-mapping of the 13q14 deletion and the identification of intragenic or small alterations undetected by FISH. Moreover, additional alterations in 2p24 (MYCN) (3 pts), 8q24 (MYC) (1 pt), 9p21 (CDKN2A2B) (1 pt), 1q21 (LMNA) (1 pt), and 6q25-26 (1 pt) regions not covered by a standard FISH assay were detected and all confirmed by FISH. Our data extend previously limited evidence that MLPA may represent a useful technique for the characterization of well-known lesions as well as the investigation of additional genomic changes in CLL.     

Cytogenetic abnormalities detected by fluorescence in situ hybridization on paraffin-embedded chronic lymphocytic leukemia/small lymphocytic lymphoma lymphoid tissue biopsy specimens

Cytogenetic fluorescence in situ hybridization (FISH) panels are a major prognostic tool in chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), but few data exist on using paraffin-embedded extramedullary tissue biopsy specimens for these purposes. Isolated whole nuclei were extracted from 20 paraffin-embedded tissue biopsy specimens with CLL/SLL and analyzed using a standard CLL FISH panel. FISH studies were successful in 18 (90%) of 20 cases, and chromosomal abnormalities were detected in 18 (100%) of the technically successful cases. Deletion 13q14.3 was most frequent (10 [56%]; isolated in 8 and with other abnormalities in 2), followed by trisomy 12 (5 [28%]), deletion 11q22.3 (4/16 [25%]), 14q32 (IGH@) translocation (3 [17%]), and deletion 17p13.1 (1/16 [6%]). One case with IGH@ translocation showed a BCL2 translocation partner. No cases showed 6q23 deletion. Results of this FISH panel performed on 42 additional peripheral blood (PB)/bone marrow (BM) CLL specimens were similar except for a significantly greater frequency of deletion 13q14.3 in combination with other aberrations. Cytogenetic FISH studies using paraffin-embedded tissue biopsy specimens in CLL/SLL had a high yield and, with 1 exception, demonstrated a profile similar to cases diagnosed in PB/BM.     

The influence of cytogenetic aberrations on gene replication in chronic lymphocytic leukemia patients

Chronic lymphocytic leukemia (CLL) is the most common leukemia in humans, with the major cytogenetic aberrations of trisomy 12 and deletion of 13q14. This study examined the influence of these aberrations on general gene replication. The study group included three subgroups: (1) 15 CLL patients, (2) 4 CLL patients with trisomy 12, (3) 3 CLL patients with deletions in 13q14. Five healthy individuals served as a control group. Monocolor fluorescence in situ hybridization (FISH) with probes for c-myc, HER-2/neu, and p53 was applied to lymphocyte nuclei for the evaluation of replication timing. Asynchronous replication (SD) rate was significantly higher in all CLL patients (P < 0.01) when compared to the control group and was even higher in the group of CLL patients with trisomy 12 and 13q14 deletion (P < 0.01). The asynchrony rate was significantly higher in cells with trisomy 12 for all three probes analyzed, compared to "healthy" cells in the same patients (P < 0.001). To conclude, in CLL patients with a chromosomal aberration such as trisomy 12 and 13q14 deletion we were able to demonstrate a high rate of asynchrony of replication. The high correlation between cells with trisomy 12 and SD pattern could reflect direct influence of the aberration on gene replication and cell cycle control.     

Deletion mapping of chromosome 13q in head and neck squamous cell carcinoma in Indian patients: correlation with prognosis of the tumour

Deletions in chromosome (chr.) 13q occur frequently in head and neck squamous cell carcinoma (HNSCC). Previous studies failed to identify common deleted regions in chr.13q, though several candidate tumour suppressor genes (TSGs) loci, e.g. BRCA2, RB1 and BRCAX have been localized in this chromosome, as well as no prognostic significance of the deletion has been reported. Thus, in the present study, deletion mapping of chr. 13q has been done in 55 primary HNSCC samples of Indian patients using 11 highly polymorphic microsatellite markers of which three were intragenic to BRCA2 gene, one intragenic to RB1 gene and another from BRCAX locus. The deletion in chr.13q was significantly associated with progression of HNSCC. High frequencies (27-39%) of loss of heterozygosity were found in 13q13.1 (BRCA2), 13q14.2 (RB1), 13q21.2-22.1 (BRCAX) and 13q31.1 regions. Deletions in the BRCA2 and RB1 regions were significantly correlated. The four highly deleted regions were associated with clinical stage and histological grades of the tumour as well as poor patient outcome. Deletion in the 13q31.1 region was only found to be associated with HPV infection. High frequencies (11-23%) of microsatellite size alteration (MA) were seen to overlap with the highly deleted regions. Forty per cent of the samples showed rare biallelic alteration whereas loss of normal copy of chromosome 13q was seen in five tumours. Thus, it seems that the putative TSGs located in the BRCAX and 13q31.1 regions as well as the BRCA2 and RB1 genes may have some cumulative effect in progression and poor prognosis of HNSCC. Significant association between deletion in BRCA2 and RB1 gene loci may indicate functional relationship between the genes in this tumour progression.     

Detection of chromosomal abnormalities associated with chronic lymphocytic leukemia: what is the best method?

B-cell chronic lymphocytic leukemia (CLL) follows a heterogeneous clinical course, for which several biological markers may predict clinical outcome. Cytogenetic aberrations are considered major prognostic indicators for predicting the survival of CLL patients. Given the difficulties in obtaining abnormal metaphases in CLL, fluorescent in situ hybridization (FISH) with specific probes is generally used to detect the most frequent abnormalities. To determine the best strategy for identifying cytogenetic abnormalities, we compared results obtained by FISH analysis on peripheral blood mononuclear cells with those obtained by FISH after culture with mitogens. We studied 46 CLL patients selected from two different institutions. The most frequent structural aberrations leading to loss of genetic material were loss of the 13q14 region, in 32 cases (70%), and loss of TP53 in 11 cases (24%). Of the 46 cases patients, 10 patients (21.7%) had deletion of the ATM locus at 11q22 and 8 patients (17%) had trisomy 12. Results could be interpreted as discordant in four cases in which the abnormal clone was small and both values were around the threshold. FISH performed on stimulated cells detected the same frequency of abnormalities as FISH performed without culture. This equivalence between the two techniques allows performing both conventional cytogenetic and FISH analyses on the same sample.     

Familial 22q11.2 deletions in DiGeorge/velocardiofacial syndrome are predominantly smaller than the commonly observed 3Mb.

PURPOSE: DiGeorge/velocardiofacial syndrome (DG/VCFS) is the most common cytogenetically characterized microdeletion of 22q11.2 region. In approximately 90% of patients, the deletion size is 3 Mb, whereas the remaining range from 1.5 to 2.5 Mb. The purpose of this study was to test the hypothesis that small deletions may be more easily tolerated in a familial fashion than larger deletions, especially for this syndrome. METHOD: Sixteen FISH probes designed from bacterial artificial chromosomes (BACs) and P1 artificial chromosomes (PACs) mapped to 22q11.2 were used to determine the deletion sizes in 22 individuals from ten families with familial 22q11.2 deletion detected by standard FISH tests. RESULT: Seven families had deletions of < 3 Mb ( approximately 1.5 Mb) in size and 3 families had the common 3-Mb deletion. The 70% frequency of smaller sized deletions among this group of patients with familial del(22)(q11.2) is significantly higher than that reported among unselected group of patients with del(22)(q11.2) (P < 0.0001, Fisher exact test). CONCLUSION: Familial del(22)(q11.2) are predominantly smaller than the common deletion size of 3 Mb, indicating that there may be some underlying mechanisms that favor parent-to-child transmission of smaller deletions in individuals with del(22)(q11.2), therefore, underscoring the need to exclude a familial basis in cases of del(22)(q11.2) smaller than 3 Mb.     

Multicolor FISH in chronic lymphocytic leukemia. An interphase study of patients with early-onset disease

Trisomy 12 and deletions of 13q14.2 and 14q32 are the most common chromosome abnormalities in patients with B-chronic lymphocytic leukemia (B-CLL), but whether specific chromosomal defects influence the course of B-CLL is still a matter of discussion. The aim of our study was to assess the possible correlation between cytogenetic findings and clinical characteristics. Thirty patients with previously untreated early-onset B-CLL were recruited. The incidence of trisomy 12, and observations of 13q14.2 and 14q32 was analyzed in unstimulated bone marrow cells by means of multicolor interphase FISH. No correlation was found between trisomy 12 and the patients' clinical characteristics. The analysis of the patients with trisomy 12 and observations of 13q14.2 and 14q32 revealed heterogeneity of the leukemic cell population, thus indicating that these chromosomal abnormalities are probably a secondary event in CLL leukemogenesis. The finding of RB1 gene nullisomy and 14q32 deletions in patients at an advanced clinical stage suggests a possible correlation between these rearrangements and disease progression. Multicolor FISH analysis in B-CLL provides important diagnostic, clinical, and prognostic information that may help in assessing prognosis and making treatment decisions.     

Intratumoral genetic heterogeneity and number of cytogenetic aberrations provide additional prognostic significance in chronic lymphocytic leukemia

PurposeChronic lymphocytic leukemia (CLL) is a heterogeneous disease with cytogenetic aberrations that are still considered the gold standard of prognostic factors. However, heterogeneity remains within each cytogenetic group, especially in patients with concomitant cytogenetic aberrations.MethodsA panel of DNA probes was used to detect cytogenetic aberrations, including RB1/D13S25 at 13q14, ATM at 11q22, TP53 at 17p13, CEP12 and IGH translocation at 14q32, by fluorescence in situ hybridization. A comprehensive method integrating the number of cytogenetic aberrations and intratumoral genetic heterogeneity was used to analyze the prognosis for patients with concomitant aberrations.ResultsWithin the conventional favorable or neutral prognostic groups (i.e., with del 13q, trisomy 12, and/or t(14q32)), the coincidence of these three aberrations worsened survival in terms of time to first therapy, progression-free survival, and overall survival. However, within the conventional unfavorable prognostic group (i.e., del 11q or del 17p), patients with a minor unfavorable clone had an unexpected survival advantage compared with patients with a major unfavorable clone. A new cytogenetic prognostic system that integrates the number of cytogenetic aberrations and intratumoral genetic subclones was more precise than the conventional system.ConclusionThe number of cytogenetic aberrations and the size of intratumoral genetic subclones should be comprehensively considered to determine the prognosis for CLL.     

13q deletions in B-cell lymphoproliferative disorders: frequent association with translocation

The 13q14 deletion is the most frequent abnormality in chronic lymphocytic leukemias/small lymphocytic lymphomas, and this early rearrangement is observed from the start of the disease. The systematic use of a panel of interphase fluorescence in situ hybridization (FISH) may not reveal some probes (targeting chromosomes 11q, 13q, 17p, and chromosome 12) structural abnormalities. In this series, we analyzed metaphases by conventional cytogenetics, followed by interphase and metaphase fluorescence in situ hybridization. We were able to observe 17 cases of 13q translocations with deletions in eight of them. Three distinct regions were involved by translocations in association with or without deletions: a region centromeric to RB1 (13q11 approximately 13), a zone telomeric to D13D25 (13q21 approximately 31), and a 13q14 region deliniated by RB1 and D13S25. In this area, the deletion was variable: RB1 alone (one case), D13S319 approximately D13S25 (five cases), and from RB1 to D13S25 (two cases). The very high frequency of 13q14 loss suggests that these deletions are of pathogenetic importance, but, the importance of the translocations remains to be determined.     

Subtle overlapping deletions in the terminal region of chromosome 6q24.2-q26: Three cases studied using FISH

Interstitial deletions in the terminal region of chromosome 6 are rare. We describe three new cases with subtle interstitial deletions in the q24-q26 region of the long arm of chromosome 6. The karyotypes were analyzed at a 550 band level. Patient1 is a 9-month-old boy with an interstitial deletion, del(6)(q24.2q25.1), developmental delay, low birth weight, hypotonia, heart murmur, respiratory distress, craniofacial and genital anomalies. This is the first report of a case with deletion del(6)(q24.2q25.1). Patient 2 is a 17-year-old young man with an interstitial deletion del(6)(q25.1q25.3), developmental delay, short stature, mental retardation, autism, head, face, chest, hand and feet anomalies and a history of seizures. For the first time autism was described as a manifestation in 6q deletions. Patient 3 is baby boy with a de novo interstitial deletion, del(6)(q25.1q26), anomalies of the brain, genital organs, limbs and feet. This is the first report of a case with deletion, del(6)(q25.1q26). In all three patients, fluorescence in situ hybridization (FISH) using chromosome 6 painting probe ruled out an insertion. The ESR (6q25.1) and TBP (6q27) probes were used to confirm the breakpoints. Since TBP signal is present in all cases, it confirmed an interstitial deletion proximal to this probe. Patient 1 has a deletion of the ESR locus; Patient 2 and 3 have signals for the ESR locus on both chromosomes 6. Therefore the deletion in Patients 2 and 3 are between ESR and TBP loci distal to that of Patient 1. FISH validated the deletion breakpoints assessed by conventional cytogenetics. Am. J. Med. Genet. 87:17–22, 1999. © 1999 Wiley-Liss, Inc.